Skip to main content Accessibility help


  • Access
  • Cited by 1


      • Send article to Kindle

        To send this article to your Kindle, first ensure is added to your Approved Personal Document E-mail List under your Personal Document Settings on the Manage Your Content and Devices page of your Amazon account. Then enter the ‘name’ part of your Kindle email address below. Find out more about sending to your Kindle. Find out more about sending to your Kindle.

        Note you can select to send to either the or variations. ‘’ emails are free but can only be sent to your device when it is connected to wi-fi. ‘’ emails can be delivered even when you are not connected to wi-fi, but note that service fees apply.

        Find out more about the Kindle Personal Document Service.

        In vitro larvicidal and in vivo anthelmintic effects of Oxalis tetraphylla (Oxalidaceae) hydroalcoholic extract against Haemonchus contortus in lambs
        Available formats

        Send article to Dropbox

        To send this article to your Dropbox account, please select one or more formats and confirm that you agree to abide by our usage policies. If this is the first time you use this feature, you will be asked to authorise Cambridge Core to connect with your <service> account. Find out more about sending content to Dropbox.

        In vitro larvicidal and in vivo anthelmintic effects of Oxalis tetraphylla (Oxalidaceae) hydroalcoholic extract against Haemonchus contortus in lambs
        Available formats

        Send article to Google Drive

        To send this article to your Google Drive account, please select one or more formats and confirm that you agree to abide by our usage policies. If this is the first time you use this feature, you will be asked to authorise Cambridge Core to connect with your <service> account. Find out more about sending content to Google Drive.

        In vitro larvicidal and in vivo anthelmintic effects of Oxalis tetraphylla (Oxalidaceae) hydroalcoholic extract against Haemonchus contortus in lambs
        Available formats
Export citation


The in vitro larvicidal and in vivo anthelmintic effects of Oxalis tetraphylla hydroalcoholic extract (HE), against Haemonchus contortus in experimentally infected lambs, were assessed. We used a microtitration plate method, comprising the following two stages. Stage 1: 20 μl of water containing 200 sheathed H. contortus infective larvae (ShHcl) were deposited in every well of three series; then, the series 2 and 3 wells were treated with 80 μl 1% ivermectin and O. tetraphylla HE at 20 mg/ml, respectively. Stage 2: the same procedure was performed replacing the ShHcl with exsheathed larvae (ExShHcl). Evaluations were performed after 24 and 48 h. The total numbers of dead and live larvae were counted. A second experiment evaluated the reduction in nematode egg populations in the faeces of lambs treated orally with the O. tetraphylla HE. The 27 lambs used were divided into Groups 1, 2 and 3 (n = 9), which were administered water (positive control), levamisole 1 m (7.5 mg/kg body weight (BW), as a unique dose) and O. tetraphylla HE (20 mg/kg BW), respectively. The plant HE was administered daily for 8 days. The in vitro assay showed 80.9% and 86.5% larval mortality of ShHcl after 24 and 48 h, respectively, while the corresponding mortality values for ExShHcl were 97 and 99%, respectively. The in vivo assay showed variability in the eggs/gram of faeces (epg) values; however, at the end of the trial, the average reduction in the epg values of the O. tetraphylla HE group was 45.6% (P < 0.05). Oxalis tetraphylla HE contains compounds that belong to the flavonol group with anthelmintic activity.


Sheep haemonchosis is considered to be one of the main health problems affecting sheep flocks worldwide (Fiaz-Qamar et al., Reference Fiaz-Qamar, Maqbool and Ahmad2011). Infected animals show clinical signs such as anorexia, weight loss, diarrhoea, submaxillary oedema, slow growth, malnutrition (Prakash & Bano, Reference Prakash and Bano2010) and even death of young animals (Garedaghi et al., Reference Garedaghi, Hashemzadefarhang and Esmaeli2013). Haemonchosis and other gastrointestinal parasitic nematodes (GIPN) have been controlled mainly with chemical anthelmintic drugs that are administered as a continuous treatment strategy for de-worming animals. However, this system has led to the development of anthelmintic resistance (AR) (Torres-Acosta et al., Reference Torres-Acosta, Mendoza-de-Gives, Aguilar-Caballero and Cuéllar-Ordaz2012) as well as health risks due to the possible persistence of chemical residues in animal products. The use of ethnoveterinary or traditional medicines in veterinary practice has gained the attention of numerous researchers worldwide, and some plants have been reported to be excellent candidates for controlling animal parasitosis caused by nematodes (Akerreta et al., Reference Akerreta, Calvo and Cavero2010; Bharati & Sharma, Reference Bharati and Sharma2012; Hassan et al., Reference Hassan, Murad, Tariq and Ahmad2014; Saha et al., Reference Saha, De Sarker and Sen2014). Plants such as Trifolium repens and Vernona anthelma are currently being investigated as possible sources of agents to control GIPN in ruminants (Heckendorn et al., Reference Heckendorn, Häring, Maurer, Senn and Hertzberg2007; Alawa et al., Reference Alawa, Adamu, Gefu, Ajanusi, Abdu and Chiezey2010). Oxalis tetraphylla is a bulbous plant from Mexico and Guatemala that belongs to the Oxalidaceae family, which has approximately 900 species (Burger, Reference Burger1991). It is widely distributed in South America, Mexico and Africa (Loudon, Reference Loudon2009). It is traditionally called ‘Trebol de la Buena Suerte’ (good luck clover) and possesses four triangular green leaves with purple colour diffused at the centre of the leaves (Brickell & Zuk, Reference Brickell and Zuk1997) (fig. 1). Although this plant has been used for human consumption, it is mostly used for cattle grazing (Romero, Reference Romero2000). The most reliable assay for assessing the in vivo anthelmintic activity of plant extracts involves slaughtering the animal and then counting the adult worms recovered from the digestive tract. However, this method requires the euthanization of numerous animals, which is very expensive. Another technique that allows the identification of potential anthelmintic compounds is based on counting the number of eggs/gram of faeces (epg) before and after treatments (Irum et al., Reference Irum, Ahmed, Mukhtar, Mushtaq, Mirza, Donskow-Łysoniewska, Qayyum and Simsek2015). The method is simple, inexpensive and avoids the euthanization of animals. The present study was designed to assess both the in vitro nematocidal activity against Haemonchus contortus infective larvae (L3) and the in vivo effect of oral administration of O. tetraphylla hydroalcoholic extract (HE) on the faecal egg count in lambs experimentally infected with H. contortus.

Fig. 1. Leaves of the leguminous plant Oxalis tetraphylla collected from the top of the Woodland Mountain at San Juan Tlacotenco village, Tepoztlán, Morelos State, Mexico.

Materials and methods

Plant material

Wild specimens of O. tetraphylla were collected from a forest at the top of the ‘Cerro light’ mountain (mountain of light) in the village of San Juan Tlacotenco, Tepoztlán Municipality, Morelos, Mexico, in January 2012. A voucher specimen of the plant material was deposited at the herbarium of the National Institute of Anthropology and History of Mexico, Cuernavaca City, Morelos State, Mexico (registration number: 2058). A biologist, Margarita Aviléz, established the taxonomy of the plant specimen.

Extract preparation

Five kilograms of the wild leaves and stems were dried and then extracted three times by maceration in an ethanol–water mixture (60:40, v/v, 15 litres; Merck, Darmstadt, Germany) for 24 h at room temperature (18–25°C). This maceration process was described previously by De Jesús Gabino et al. (Reference De Jesús-Gabino, Mendoza de Gives, Salinas-Sánchez, López-Arellano, Liébano-Hernández, Hernández-Velázquez and Valladares-Cisneros2010). The HE of each plant material sample was concentrated and completely dried using a rotatory evaporator (Heidolph Laborota 4000; Heidolph Instruments, Schwabach, Germany) under reduced pressure at 50–60°C to obtain 38.1 g of HE extract.

Phytochemical analysis using thin layer chromatography

To identify the chemical compounds in the O. tetraphylla HE, we used a thin layer chromatography (TLC) technique to analyse specific chemical reactions (Wagner et al., Reference Wagner, Bladt and Zgainski1984). We used a visualization agent (2-aminoethyl diphenylborinate in polyethylene glycol) to detect flavonoids, which were observed under ultraviolet (UV) light at a long wavelength (360 nm). A comparison of the TLC analysis of samples previously isolated indicated that the detected compounds were β-sitosterol and stigmasterol sterols, which are known to have widespread occurrence in the plant kingdom. The compounds were revealed using the Komarowski reagent (4-hydroxybenzaldehyde in sulphuric acid).

Phytochemical analysis using high-performance liquid chromatography

The high-performance liquid chromatography (HPLC) analysis was performed using a Waters 2695 separation module system equipped with a Waters 2996 photodiode array detector and the Empower Chromatographic Manager version 1 software (Waters, Milford, Connecticut, USA). The analysis was performed using a Merck Superspher® RP-18 (Merck) column (5 μm, 100 mm). The mobile phase consisted of a gradient system of water, trifluoroacetic acid (TFA 0.5%, solvent A) and acetonitrile (solvent B). The chromatographic process was run for 28 min on the following schedule: 1–2 min (solvent A:B, 100:0), 3–4 min (90:10), 5–7 min (80:20), 8–14 min (70:30), 15–18 min (60:40), 19–22 min (20:80), 23–26 min (0:100) and 27–28 min (100:0). The flow rate was 1 ml/min while the sample was injected in a volume of 10 μl and at a 1 ml/min flow rate. The detection wavelength was 190–400 nm.

Pharmacological evaluation

Two experiments were performed. Experiment 1 was aimed at evaluating the in vitro larvicidal activity of the O. tetraphylla HE against the L3. The second experiment evaluated the effect of oral administration of O. tetraphylla HE to lambs on the reduction of numbers of H. contortus eggs eliminated in the faeces.

Biological material

L3 processing

Faecal samples of sheep experimentally infected with H. contortus were processed by preparing faecal cultures and extracting the infective larvae using a Baermann's funnel. The L3 were washed several times using density gradients of a 40% saccharose solution, rinsed and then suspended in sterile water.

Experiment 1: in vitro assessment of effects of O. tetraphylla HE against L3

Experimental design

Two experimental stages were used to assess the in vitro activity of the O. tetraphylla HE against sheathed and exsheathed L3. The larvae/extract treatment was carried out in 96-well plates. Three wells per treatment were considered as three experimental units. The design of each experimental stage was similarly structured, and three series of three wells were treated as follows. Stage 1: first, 20 μl of water containing 200 sheathed L3 were placed in every well of the three series. Series 1 contained only larvae; in series 2 and 3, 80 μl of 1% ivermectin or O. tetraphylla HE at a concentration of 20 mg/ml were added and mixed. Stage 2 followed the same procedure as that used in Stage 1 except that exsheathed larvae were used.

Two incubation periods were used (24 and 48 h post-treatment). After incubation, ten 5-μl aliquots (considered as n = 10 aliquots/well from three experimental units per treatment) were placed on a slide and observed under a microscope at 4× and 10× magnifications. The total, dead and live larvae were counted. The criteria for identifying the dead and live larvae were based on their mobility/immobility; physical stimuli were applied to confirm the dead and live larvae. The proportions of dead and live larvae were estimated for each series. The proportion of live larvae in series 1 (control, water) was considered as 100% and used for comparison. Ponder adjustments were performed when necessary considering the number of larvae that died for reasons other than the treatments.

Statistical analysis

The data were transformed (√x + 0.5) using a completely random design. The means of the live larvae were compared using an analysis of variance (ANOVA), and the complementary Tukey's test was performed to identify the differences between treatments. The statistical analysis software (SAS) program (version 8) was used (SAS Institute Inc., Cary, North Carolina, USA). The percentage in vitro efficacy of the extract was estimated using the following formula (Eguale & Giday, Reference Eguale and Giday2009):

$$\% {\rm Efficacy} = \displaystyle{{\matrix{{\bar {\rm x}} \; {\hbox {live larvae in control group}} \cr - {\bar {\rm x}}\; {\hbox {live larvae in treated group}}}} \over {{\bar {\rm x}} \; {\hbox {live larvae in control group}}}} \times 100$$

Experiment 2: evaluation of the effect of oral administration of O. tetraphylla HE on the number of H. contortus eggs eliminated in sheep faeces

Experimental groups of animals

Twenty-seven Pelibuey lambs aged 4–6 months, previously infected with 350 L3/kg body weight (BW), were randomly assigned to three groups of nine lambs each. Group 1 was treated with water (positive control); Group 2, levamisole, 7.5 mg/kg BW (one single dose); and Group 3, O. tetraphylla HE, 20 mg/kg BW (orally administered daily for 8 days). The animals were maintained in individual paddocks and received a dried alfalfa nutritional regime and water ad libitum. The experimental groups were established based on their epg counts on day −3 and treatments were administered on day 0.

Faecal sampling

Faecal samples were collected directly from the rectum of each lamb on days −3, 0, 2, 4, 7, 9 and 11 of the experiment. The average number of nematode eggs eliminated (epg) by each lamb in every group was estimated using the McMaster technique (Paul et al., Reference Paul, Torgerson, Höglund and Furrer2014).

Statistical analysis

The data were analysed using the SAS program. The epg values were 10log transformed (epg + 1) to achieve a normal distribution approximation (Bouix et al., Reference Bouix, Krupinski, Rzepecki and Nowosad1998) and an analysis of repeated measures over time series was performed. A Duncan's multiple range test was used to compare the means, and the following statistical model was used:

$$Y_{ijkl} = {\rm \mu} {\rm} + E_i + T_j + {\rm} (E \times T)_{ij} + A_k + {\rm \varepsilon} _{ijk}$$

where Y ijkl = variable/response (epg), μ = general mean, E i = ith effect/sample day (i = 11), T j = jth effect/treatment (j = 4), (E × T)ij = interaction effect between treatments/sampling days, A k = random effect of the kth lambs, εijk = experimental error.


Experiment 1

In the in vitro assay, the proportions of dead and total larvae (sheathed and exsheathed) recovered after incubation for 24 and 48 h with the different treatments are shown in tables 1 and 2, respectively. After a 24-h incubation, the mortality rate of the sheathed larvae in series 1 (control, water) was low and no mortality was observed for exsheathed larvae. In series 2 (control, ivermectin), no live sheathed or exsheathed larvae were observed. However, when the mortality of sheathed larvae was compared with that of its control (water-treated), a 97.6% ponderate mortality (real mortality) was recorded. In series 3, (O. tetraphylla HE), 80.9% and 97% ponderate mortalities were observed for the sheathed and exsheathed larvae, respectively (table 1). After a 48-h incubation, a low mortality was observed in the sheathed larvae of the control (series 1). No mortality was observed in the exsheathed larvae of the control series. Series 2 (control, ivermectin) exhibited a 100% mortality in both sheathed and exsheathed larvae, while series 3 (O. tetraphylla HE) showed corresponding ponderate mortalities of 86.87 and 98.85%, respectively (table 2).

Table 1. Proportion of dead and total Haemonchus contortus infective larvae (L3, sheathed and exsheathed) exposed to Oxalis tetraphylla hydroalcoholic extract (HE) after 24-h incubation, and mortality percentages.

n = 3 (3 wells, 10 aliquots per well). P < 0.05.

Table 2. Proportion of dead and total Haemonchus contortus infective larvae (L3, sheathed and exsheathed) exposed to Oxalis tetraphylla hydroalcoholic extract (HE) after 48-h incubation, and mortality percentages.

n = 3 (3 wells, 10 aliquots per well). P < 0.05.

Experiment 2

The results of experiment 2, including the mean number of H. contortus eggs eliminated per group of animals and the egg reduction percentage in the different groups relative to the control group are shown in fig. 2. The overall results of group 1 (control, water) showed the highest epg values. In group 2 (levamisole), no eggs were recorded in the faecal samples, indicating a 100% efficacy, while group 3 (O. tetraphylla HE) showed variable reduction percentages ranging between 23.1 and 63.7%. The maximum values were obtained on day 11 after treatment. The overall reduction percentage at the end of the experiment for the O. tetraphylla HE was 45.6% (fig. 2, P < 0.05). Statistical differences were found between the control group treated with water (positive control) without any de-worming treatment and the extract-treated group (P < 0.05). However, this group exhibited a lower effect than that of the levamisole-treated group (fig. 2).

Fig. 2. Haemonchus contortus egg counts per gram faeces from lambs receiving the three different treatments.

Analysis of phytochemicals

The phytochemical analysis of the O. tetraphylla HE showed a positive reaction to the reagent that revealed the presence of classic flavonoids and sterols. Figure 3 shows the HPLC chromatogram generated at 345 nm, indicating a mixture of compounds corresponding to flavonols according to UV absorption (218, 272 and 357 nm).

Fig. 3. Chromatogram generated from an Oxalis tetraphylla hydroalcoholic extract showing several compounds identified at different times. Compounds were eluted between 2 and 12 min. Compounds reaching values higher than 350 nm corresponded to flavonoids and sterols.


Plants contain numerous beneficial nutritional compounds and are used as an important source of protein, starch, minerals and vitamins. Furthermore, they are also considered to contain medicinal compounds. including phenols, inositol phosphates and oligosaccharides, that are beneficial to cattle and small ruminants (Schuster-Gajzágó, Reference Schuster-Gajzágó and Fuleky2004). To date, very little information is available about the medicinal effects of the plant, O. tetraphylla (Oxalidaceae). The results of the present study showed the potent in vitro nematocidal activity of O. tetraphylla HE against both sheathed and exsheathed L3 after 24- and 48-h incubations. These results encouraged us to further investigate the possible anthelmintic effect of the O. tetraphylla HE in sheep infected with H. contortus.

This study provided evidence of the presence of bioactive compounds with anthelmintic activity in O. tetraphylla. Specifically, oral administration of the HE to H. contortus-parasitized sheep reduced the epg values to approximately 50% after daily treatment for 8 days. This level of reduction is considered inadequate by the World Association for the Advancement of Veterinary Parasitology (WAAVP) (Coles et al., Reference Coles, Jackson, Pomroy, Prichard, Himmelstjerna, Silvestre, Taylor and Vercruysse2006). Nevertheless, it is important to consider that this low epg reduction was obtained with a crude extract, and further purification could lead to the identification of a more potent molecule with improved effects. Our extensive search for relevant information did not reveal any reports of the possible ovicidal effects of O. tetraphylla extracts against any nematode. Therefore, this could be the first report on this activity in O. tetraphylla.

Furthermore, the hypothesis that this plant extract has a lethal effect against adult parasites in sheep following oral administration into the abomasum was proven. The common assay to determine the precise antiparasitic effect of plant extracts or chemical compounds involves collecting, counting and comparing the number of adult parasites in untreated (control) and treated sheep at necropsy. However, this method is expensive and requires the euthanization of the animals, as previously mentioned. Therefore, we assessed the treatment effects by determining and comparing the epg faecal reduction in treated and control groups (untreated sheep).

The low reduction in the epg values could be attributable to the use of a crude plant extract that contained numerous bioactive compounds that could interfere with the biological activity of the extract against parasites. The chromatographic purification of the extract could possibly lead to the identification of molecules with higher efficiency against the parasites than that of the crude extract. The epg reduction percentage of the O. tetraphylla HE (< 50%) was not as expected; however, we believe that purifying the plant extract could lead to the isolation of a component with a higher epg reduction percentage. Furthermore, this compound could be a useful tool in an integrated control programme including other alternative options, focused on reducing the parasitic burden through different mechanisms. For example, strategies such as the nutritional strategy of improving the quality and quantity of protein and metabolizable energy levels consumed (Yap et al., Reference Yap, Utzinger, Hattendorf and Steinman2014), and management measures such as rotational grazing (Burggraaf et al., Reference Burggraaf, Boom, Sheath and Brooky2009; Benson, Reference Benson2012) or alternated grazing with different host species (Marshall et al., Reference Marshall, Gebrelul, Gray and Ghebreiyessus2012), are beneficial. Furthermore, the use of natural nematode enemies such as nematophagous fungi of the species Duddingtonia flagrans, vaccines (Roberts et al., Reference Roberts, Antonopoulos, Haslam, Dicker, McNeilly, Johnson, Dell, Knox and Britton2013; Fawzi et al., Reference Fawzi, González-Sánchez, Corral, Cuquerella and Alunda2014), genetic selection of animals resistant to parasites (Hutchings et al., Reference Hutchings, Knowler, McAnulty and McEwan2007; Periasamy et al., Reference Periasamy, Pichler, Poli, Cristel, Cetra, Medus, Basar, Thiruvenkadan, Ramasamy, Ellahi, Mohamed, Teneva, Shamsuddin, García and Diallo2014), and copper particles (Sayward & Sayward, Reference Sayward and Sayward2014) are other promising measures. In addition, other groups of plants have also been explored under different conditions to identify additional natural alternatives for control, and different results have been published (table 3).

Table 3. In vitro and in vivo effect of different plants against sheep gastrointestinal parasitic nematodes.

In conclusion, the present study provides relevant information about the presence of anthelmintic compounds in the crude HE of the bulbous plant, O. tetraphylla. Furthermore, the type of flavonoids identified killed the infective stages of H. contortus in in vitro assays and reduced the H. contortus egg nematode population in the faeces of treated animals. The results of this study could be used as a reference for further investigations focused on identifying a biomolecule with anthelmintic activity from the bulbous plant, O. tetraphylla, for the control of sheep haemonchosis.


This research formed part of the theses of B.J.G.-C. and M.R.-L., submitted towards the qualification of Biotechnologist Engineers Labastida, under the direction of P.M. de G. at the Universidad Politécnica del Estado de Morelos, Jiutepec, Morelos, Mexico.

Financial support

The present research received financial support from Instituto Nacional de Investigaciones Forestales, Agricolas y Pecuarias (INIFAP)-México (Fondos Fiscales-2012) from the Project ‘Assessment of four plant extracts as phytotherapeutics against sheep parasitic nematodes’.

Conflict of interest


Ethical standards

The sheep were strictly maintained under the Norma Oficial Mexicana (Official Rule Number) NOM-051-ZOO-1995 ( and the LEY Federal de Sanidad Animal (Federal law for animal health) DOF 07-06-2012 ( These guidelines specify that all the procedures performed in studies involving animals must follow the Federal Law and Official Rule strictly in accordance with the ethical standards of INIFAP. Furthermore, the guidelines are based, in part, on the Guide for the care and use of laboratory animals published by the Institute of Laboratory Animals Resources Commission on Life Sciences, National Research Council, 1996.


Akerreta, S., Calvo, M.I. & Cavero, R.Y. (2010) Ethnoveterinary knowledge in Navarra (Iberian Peninsula). Journal of Ethnopharmacology 130, 369378.
Alawa, C.B.I., Adamu, A.M., Gefu, J.O., Ajanusi, O.J., Abdu, P.A. & Chiezey, N.P. (2010) In vivo efficacy of Vernonia amigdalina (Compositae) against natural helminth infection in Bunaji (Bos indicus) calves. Pakistan Veterinary Journal 30, 215218.
Ansa, P., Athul, P.V., Charan, A. & Afeefa, T.P. (2009) Anthelmintic activity of Macrotyloma uniflorm. Hygeia Journal for Drugs and Medicines 1, 2627.
Benson, H. (2012) Parasite loads of beef cattle in a rotationally grazed closed herd. MSc degree dissertation thesis, Murray State University, Kentucky, USA.
Bharati, A.K. & Sharma, B.L. (2012) Plants used as ethnoveterinary medicines in Sikkim Himalayas. Ethnobotany Research and Applications 10, 339356.
Bouix, J., Krupinski, J., Rzepecki, R. & Nowosad, B. (1998) Genetic resistance to gastrointestinal nematode parasites in Polish long-wool sheep. International Journal for Parasitology 28, 17971804.
Brickell, C. & Zuk, J.D. (1997) The American Horticultural Society A–Z encyclopedia of garden plants. New York, USA, DK Publishing.
Burger, W. (1991) Flora Costaricensis. Fieldiana, Botany. New Series, No. 28. Publication 1428. Chicago, Illinois, USA, Field Museum of Natural History.
Burggraaf, V.T., Boom, C.J., Sheath, G.W. & Brooky, A.R. (2009) The effect of rotational grazing by either calves, cows, lambs or ewes on the removal of herbage contaminated with gastro-intestinal parasite larvae around cattle faecal pats, New Zealand. Journal of Agricultural Research 52, 289297.
Coles, G.C., Jackson, F., Pomroy, W.E., Prichard, R.K., Himmelstjerna, V.S., Silvestre, A., Taylor, M.A. & Vercruysse, J. (2006) The detection of anthelmintic resistance in nematodes of veterinary importance. Veterinary Parasitology 136, 168185.
De Jesús-Gabino, A.F., Mendoza de Gives, P., Salinas-Sánchez, D.O., López-Arellano, M.E., Liébano-Hernández, E., Hernández-Velázquez, V.M. & Valladares-Cisneros, G. (2010) Anthelmintic effects of Prosopis laevigata n-hexanic extract against Haemonchus contortus in artificially infected gerbils (Meriones unguiculatus). Journal of Helminthology 84, 7175.
Eguale, T. & Giday, M. (2009) In vitro anthelmintic activity of three medicinal plants against Haemonchus contortus. International Journal of Green Pharmacy 3, 2934.
Fawzi, E.M., González-Sánchez, M.E., Corral, M.J., Cuquerella, M. & Alunda, J.M. (2014) Vaccination of lambs against Haemonchus contortus infection with a somatic protein (Hc23) from adult helminths. International Journal of Parasitology 44, 429436.
Fiaz-Qamar, M., Maqbool, A. & Ahmad, N. (2011) Economic looses due to haemonchosis in sheep and goats. Science International (Lahore) 23, 321324.
Garedaghi, Y., Hashemzadefarhang, H. & Esmaeli, A. (2013) Study on the prevalence and species composition of abomasal nematodes in small ruminants slaughtered at Behshahr Town. Iranian Journal of Veterinary Advance 3, 5559.
Hassan, H.U., Murad, W., Tariq, A. & Ahmad, A. (2014) Ethnoveterinary study of medicinal plants in Malakand Valley, District Dir (Lower), Khyber Pakhtunkhwa, Pakistan. Irish Veterinary Journal 67, 16.
Heckendorn, F., Häring, A.D., Maurer, V., Senn, M. & Hertzberg, H. (2007) Individual administration of three tanniferous forage plants to lambs artificially infected with Haemonchus contortus and Cooperia curticei. Veterinary Parasitology 146, 123134.
Hutchings, M.R., Knowler, K.J., McAnulty, R. & McEwan, J.C. (2007) Genetically resistant sheep avoid parasites to a greater extent than do susceptible sheep. Proceedings of the Royal Society of London, B 274, 18391844.
Iqbal, Z., Latee, M. & Gilani, A.H. (2010) In vivo anthelmintic activity of Azadirachta indica A. Juss seeds against gastrointestinal nematodes of sheep. Veterinary Parasitology 168, 342345.
Irum, S., Ahmed, H., Mukhtar, M., Mushtaq, M., Mirza, B., Donskow-Łysoniewska, K., Qayyum, M. & Simsek, S. (2015) Anthelmintic activity of Artemisia vestita Wall ex DC. and Artemisia maritima L. against Haemonchus contortus from sheep. Veterinary Parasitology 15, 451455.
Jabbar, A., Zaman, A.M., Iqbal, Z., Yaseen, M. & Shamim, A. (2007) Anthelmintic activity of Chenopodium album (L.) and Caesalpinia crista (L.) against trichostrongylid nematodes of sheep. Journal of Ethnopharmacology 114, 8691.
Lange, K.C., Olcott, D.D., Miller, J.E., Mosjidis, J.A., Terril, T.H., Burke, J.M. & Kearney, M.T. (2006) Effect of Sericea lespedeza (Lespedeza cuneata) fed as hay on natural and experimental Haemonchus contortus infections in lambs. Veterinary Parasitology 141, 273278.
Loudon, J. (2009) The horticulturist. 707 pp. London, Applewood Books.
Marshall, R., Gebrelul, S., Gray, L. & Ghebreiyessus, Y. (2012) Mixed species grazing of cattle and goats on gastrointestinal infections of Haemonchus contortus. American Journal of Animal Veterinary Science 7, 6166.
Naumann, H.D., Armstrong, S.A., Lambert, B.D., Muira, J.P., Tedeschid, L.O. & Kothmann, M.M. (2014) Effect of molecular weight and concentration of legume condensed tannins on in vitro larval migration inhibition of Haemonchus contortus. Veterinary Parasitology 199, 9398.
Paul, M., Torgerson, R.P., Höglund, J. & Furrer, R. (2014) Hierarchical modelling of faecal egg counts to assess anthelmintic efficacy. Journal of the Royal Statistical Society. Available at “arXiv (stat.AP)” (accessed 12 January 2014).
Periasamy, K., Pichler, R., Poli, M., Cristel, S., Cetra, B., Medus, D., Basar, M., Thiruvenkadan, A.K., Ramasamy, S., Ellahi, B.M., Mohamed, B., Teneva, A., Shamsuddin, M., García, P.M. & Diallo, A. (2014) Candidate gene approach for parasite resistance in sheep – variation in immune pathway genes and association with fecal egg count. PLoS One 9, e88337.
Pollard, D. (2009) Dose titration of Sericea lespedeza leaf meal on Haemonchus contortus infection in crossbred lambs. Doctoral dissertation, Faculty of the Louisiana State University and Agricultural and Mechanical College, USA
Prakash, V. & Bano, S. (2010) Symptoms and pathogenicity of Haemonchus contortus in sheep. Asian Journal of Animal Science 4,186187.
Roberts, B., Antonopoulos, A., Haslam, S.M., Dicker, A.J., McNeilly, T.N., Johnson, S.L., Dell, A., Knox, D.P. & Britton, C. (2013) Novel expression of Haemonchus contortus vaccine candidate aminopeptidase H11 using the free-living nematode Caenorhabditis elegans. Veterinary Research 44, 115.
Romero, L.C.E. (2000) Influencia del pastoreo en la concentración de fenoles totales y taninos condensados en Gliricidia sepium en el trópico seco. Livestock Research Rural Development 12, 410.
Saha, R.M., De Sarker, D. & Sen, A. (2014) Ethnoveternary practices among the tribal community of Malda district of West Bengal, India. Indian Journal of Traditional Knowledge 13, 359367.
Sayward, G. & Sayward, L. (2014) Effect of copper on both wool color and parasite load of Icelandic sheep. Ying Tri-Regional Science Fair 23 March 2014. Available at (accessed 23 March 2014).
Schuster-Gajzágó, I. (2004) Nutritional aspects of legumes. pp. 101114 in Fuleky, G. (Ed.) Cultivated plants, primarily as food sources, Vol 1. Encyclopedia of Life Support System (EOLSS), developed under the auspices of the UNESCO. Oxford, UK, Eolss Publishers.
Torres-Acosta, J.F.J., Mendoza-de-Gives, P., Aguilar-Caballero, A.J. & Cuéllar-Ordaz, J.A. (2012) Anthelmintic resistance in sheep farms: Update of the situation in the American continent. Veterinary Parasitology 189, 8996.
Von Son-de Fernex, E., Alonso-Díaz, M.A., Valles-de la Mora, B. & Capetillo-Leal, C.M. (2012) In vitro anthelmintic activity of five tropical legumes on the exsheathment and motility of Haemonchus contortus infective larvae. Experimental Parasitology 131, 413418.
Wagner, H., Bladt, S. & Zgainski, E.M. (1984) Plant drug analysis. Berlin, Germany, Springer-Verlag.
Yap, P., Utzinger, J., Hattendorf, J. & Steinman, P. (2014) Influence of nutrition on infection and re-infection with soil-transmitted helminths: a systematic review. Parasite & Vectors 7, 229.