Comparative anthelmintic efficacy of Arundo donax, Areca catechu, and Ferula assa-foetida against Haemonchus contortus

Eficácia anthelmíntica comparativa de Arundo donax, Areca catechu e Ferula assa-foetida contra Haemonchus contortus

Syed Nadeem Badar Zafar Iqbal Muhammad Sohail Sajid Hafiz Muhammad Rizwan Muhammad Shareef Muhammad Abdullah Malik Muhammad Nisar Khan About the authors

Abstract

In the present study, anthelmintic activities of Arundo (A.) donax L., Areca (Ar.) catechu L., and Ferula (F.) assa-foetida L. were determined. Leaves of A. donax L., latex of F. assa-foetida L. and seeds of Ar. catechu L. in different solvent fractions were subjected to in vitro (egg hatch assay; EHA, and adult motility assay; AMA) and in vivo (faecal egg count reduction test; FECRT) tests of anthelmintic activity using Haemonchus contortus model. In the AMA, crude aqueous methanol extracts (CAME) and ethyl acetate fractions of F. assa-foetida at 10 hr post-treatment showed maximum mortality of H. contortus at 12.5-50 mg mL-1. In the EHA, CAME of F. assa-foetida was identified as a potent ovicide based on its low LC50 (16.9 µg mL-1), followed in order by Ar. catechu and A. donax. Results from the FECRT also showed the extract of F. assa-foetida L. to be more effective than those of Ar. catechu L. and A. donax L., against the gastrointestinal parasitic nematodes. Chloroform and ethyl acetate fractions showed better anthelmintic activities against the adult worms in vitro, while CAME of these plants were better than their crude powders in vivo. It is recommended to document and investigate indigenous knowledge of possible medicinal plants to plan scientific trials that may justify their endorsement.

Keywords:
Arundo donax L.; Areca catechu L.; Ferula assa-foetida L.; Haemonchus contortus; in vivo; in vitro

Resumo

No presente estudo, as atividades anti-helmínticas de Arundo (A.) donax L., Areca (Ar.) Catechu L. e Ferula (F.) assa-foetida L. foram determinadas. Folhas de A. donax L., látex de F. assa-foetida L. e sementes de Ar. catechu L. em diferentes frações de solvente foram submetidos a testes in vitro (teste de eclosão de ovos, EHA e ensaio de motilidade em adultos, AMA); e in vivo (teste de redução da contagem de ovos fecais, FECRT) de atividade anti-helmíntica, usando-se Haemonchus contortus. Na AMA, extratos aquosos brutos de metanol (CAME) e frações de acetato de etila de F. assa-foetida. Dez horas pós-tratamento, apresentaram mortalidade máxima de H. contortus em 12,5-50 mg mL-1. No EHA, CAME de F. assa-foetida foi identificado como um ovicida potente baseado em seu baixo LC50 (16,9 µg mL-1), seguido em ordem por Ar. catechu e A. donax. Os resultados do FECRT também mostraram que o extrato de F. assa-foetida L. é mais eficaz do que o de Ar. catechu L. e A. donax L., contra nematoides parasitas gastrointestinais. As frações clorofórmio e acetato de etila mostraram melhores atividades anti-helmínticas contra vermes adultos in vitro, enquanto o CAME dessas plantas foi melhor do que o pó bruto in vivo. Recomenda-se documentar e investigar o conhecimento indígena de possíveis plantas medicinais para planejar ensaios científicos que possam justificar seu endosso.

Palavras-chave:
Arundo donax L.; Areca catechu L.; Ferula assa-foetida L.; Haemonchus contortus; in vivo; in vitro

Introduction

Production of the livestock depends upon regular supplies of food, healthcare and proper husbandry practices. Changes in the climate and other factors affect the prevalence of parasitic infections and consequently the productivity, longevity, fertility, and survivorship of the livestock (Thornton et al., 2009Thornton PK, van de Steeg J, Notenbaert A, Herrero M. The impacts of climate change on livestock and livestock systems in developing countries: a review of what we know and what we need to know. Agric Syst 2009; 101(3): 113-127. http://dx.doi.org/10.1016/j.agsy.2009.05.002.
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). In small-holder dairy farming systems, gastrointestinal (GI) parasites are among the major threats to animal health and welfare and have high economic significance, especially in the resource-poor countries like Pakistan (Iqbal et al., 2007Iqbal Z, Sarwar M, Jabbar A, Ahmed S, Nisa M, Sajid MS, et al. Direct and indirect anthelmintic effects of condensed tannins in sheep. Vet Parasitol 2007; 144(1-2): 125-131. http://dx.doi.org/10.1016/j.vetpar.2006.09.035. PMid:17097807.
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, Rizwan et al., 2017Rizwan HM, Sajid MS, Iqbal Z, Saqib M. Point prevalence of gastrointestinal parasites of domestic sheep (Ovis aries) in district Sialkot, Punjab, Pakistan. J Anim Plant Sci 2017; 27(3): 803-808., 2019Rizwan HM, Sajid MS, Iqbal Z, Saqib M. Association of phytomineral with gastrointestinal parasites of grazing sheep in Sialkot district, Punjab, Pakistan. Pak J Agric Sci 2019; 56(2): 459-468.; Ahmad et al., 2020Ahmad S, Sajid MS, Khan MK, Nadeem R, Rizwan HM, Malik MA, et al. Effect of trace element supplementation on the gastrointestinal parasites of grazing sheep of Multan district, Pakistan. J Anim Plant Sci 2020; 30(1): 72-80.). Lack of proper awareness, underfeeding, abundant vector populations, and availability of a wide variety of hosts provide the best opportunity for the transmission of parasitic helminths (Zvinorova et al., 2016Zvinorova PI, Halimani TE, Muchadeyi FC, Matika O, Riggio V, Dzama K. Prevalence and risk factors of gastrointestinal parasitic infections in goats in low-input low-output farming systems in Zimbabwe. Small Rumin Res 2016; 143: 75-83. http://dx.doi.org/10.1016/j.smallrumres.2016.09.005. PMid:27766016.
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).

Although most veterinarians and farmers use anthelmintic drugs for the control of helminths as a cornerstone in the small-holder dairy farming systems in the resource-poor countries like Pakistan (Akhter et al., 2014Akhter N, Arijo AG, Phulan MS, Iqbal Z. In vivo and in vitro studies on the efficacy of anthelmintics against Haemonchus contortus in goats. Pak Vet J 2014; 34(3): 329-332., Khan et al., 2017Khan MN, Sajid MS, Rizwan HM, Qudoos A, Abbas RZ, Riaz M, et al. Comparative efficacy of six anthelmintic treatments against natural infection of Fasciola species in sheep. Pak Vet J 2017; 37(1): 65-68.), the development of widespread anthelmintic resistance (Jabbar et al., 2006Jabbar A, Iqbal Z, Kerboeuf D, Muhammad G, Khan MN, Afaq M. Anthelmintic resistance: the state of play revisited. Life Sci 2006; 79(26): 2413-2431. http://dx.doi.org/10.1016/j.lfs.2006.08.010. PMid:16979192.
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), and concerns about toxicity and residues in products (Gasbarre et al., 2001Gasbarre CL, Stout LW, Leighton AE. Gastrointestinal nematodes of cattle in the northeastern US: results of a producer survey. Vet Parasitol 2001; 101(1): 29-44. http://dx.doi.org/10.1016/S0304-4017(01)00496-4. PMid:11587831.
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) have led to the revival of awareness about the potential importance of traditional veterinary practices and the need to validate their utility (Ketzis et al., 2002Ketzis JK, Taylor A, Bowman DD, Brown DL, Warnick LD, Erb HN. Chenopodium ambrosioides and its essential oil as treatments for Haemonchus contortus and mixed adult–nematode infections in goats. Small Rumin Res 2002; 44(3): 193-200. http://dx.doi.org/10.1016/S0921-4488(02)00047-0.
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, Rizwan et al., 2019Rizwan HM, Sajid MS, Iqbal Z, Saqib M. Association of phytomineral with gastrointestinal parasites of grazing sheep in Sialkot district, Punjab, Pakistan. Pak J Agric Sci 2019; 56(2): 459-468.). Ethnoveterinary medicine (EVM) can contribute to sustainable practices in the modern world, especially in developing countries (Confessor et al., 2009Confessor MVA, Mendonça LET, Mourão JS, Alves RRN. Animals to heal animals: ethnoveterinary practices in semi-arid region, Northeastern Brazil. J Ethnobiol Ethnomed 2009; 5(1): 37. http://dx.doi.org/10.1186/1746-4269-5-37. PMid:19941663.
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). Clinically useful efficacy, greater accessibility, and lower costs are the major benefits associated with EVM (Mwale et al., 2005Mwale M, Bhebhe E, Chimonyo M, Halimani TE. Use of herbal plants in poultry health management in the Mushagashe small-scale commercial farming area in Zimbabwe. Int J Appl Res Vet Med 2005; 3(2): 163-170.; Badar et al., 2017Badar N, Iqbal Z, Sajid MS, Rizwan HM, Jabbar A, Babar W, et al. Documentation of ethnoveterinary practices in district Jhang, Pakistan. J Anim Plant Sci 2017; 27(2): 398-406.; Qudoos et al., 2017Qudoos A, Khan MN, Sajid MS, Muhammad G. Correlation of trace mineral profiles with gastrointestinal worm burden in rangeland sheep of Chakwal District, Punjab, Pakistan. Int J Agric Biol 2017; 19(01): 140-144. http://dx.doi.org/10.17957/IJAB/15.0255.
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). Consequently, surveys about documentation and scientific studies on the validation of the use of plants against parasitic infections have been conducted around the world (Bizimenyera et al., 2006Bizimenyera ES, Githiori JB, Eloff JN, Swan GE. In vitro activity of Peltophorum africanum Sond. (Fabaceae) extracts on the egg hatching and larval development of the parasitic nematode Trichostrongylus colubriformis. Vet Parasitol 2006; 142(3-4): 336-343. http://dx.doi.org/10.1016/j.vetpar.2006.06.013. PMid:16899339.
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; McGaw et al., 2007McGaw LJ, Van der Merwe D, Eloff JN. In vitro anthelmintic, antibacterial and cytotoxic effects of extracts from plants used in South African ethnoveterinary medicine. Vet J 2007; 173(2): 366-372. http://dx.doi.org/10.1016/j.tvjl.2005.09.004. PMid:16239113.
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; Kareparamban et al., 2012Kareparamban JA, Nikam PH, Jadhav AP, Kadam VJ. Ferula foetida “Hing”: a review. Res J Pharm Biol Chem Sci 2012; 3(2): 775-786.; Faruque et al., 2018Faruque MO, Uddin SB, Barlow JW, Hu S, Dong S, Cai Q, et al. Quantitative ethnobotany of medicinal plants used by indigenous communities in the Bandarban district of Bangladesh. Front Pharmacol 2018; 9: 40. http://dx.doi.org/10.3389/fphar.2018.00040. PMid:29467652.
http://dx.doi.org/10.3389/fphar.2018.000...
).

The in vitro and in vivo anthelmintic efficacy of crude aqueous-methanol extracts (CAME) of Arundo (A.) donax and its various fractions (petroleum spirit, chloroform, and ethyle acetate) were reported against Haemonchus (H.) contortus (Sharatkumar et al., 2004Sharatkumar S, Dhanachand C, Mohilal N. Study on the efficacy of certain medicinal plants on gastrointestinal helminths of cattle. Indian Vet J 2004; 81(5): 497-498.; Al-Snafi, 2015Al-Snafi AE. The constituents and biological effects of Arundo donax- a review. Int J Phytopharm Res 2015; 6(1): 34-40.). Miles et al. (1993)Miles DH, Tunsuwan K, Chittawong V, Kokpol U, Choudhary MI, Clardy J. Boll weevil antifeedants from Arundo donax. Phytochemistry 1993; 34(5): 1277-1279. http://dx.doi.org/10.1016/0031-9422(91)80015-S.
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reported anthelmintic efficacy of A. donax extracts against Oesophagostomum sp. Ascaris sp., and Paramphistomum sp. of cattle. The anthelmintic efficacy of an ethanolic extract of Ferula (F.) assa-foetida against Fasciola gigantica was reported by Kumar & Singh (2014)Kumar P, Singh DK. In vitro anthelmintic activity of Allium sativum, Ferula asafoetida, Syzygium aromaticum and their active components against Fasciola gigantica. J Biol Earth Sci 2014; 4(1): 57-65.. An ethanolic extract of Areca (Ar.) catechu significantly inhibited fumarate reductase and succinate dehydrogenase activities in Cotylophoron cotylophorum (Dhanraj & Veerakumari, 2016Dhanraj KM, Veerakumari L. Effect of ethanol extract of Areca catechu on fumarate reductase and succinate dehydrogenase of Cotylophoron cotylophorum. Int J Res Develop Pharm Development Life Sci 2016;5(3): 2117-2123.). The data regarding scientific evaluation of A. donax against H. contortus models is available; however, we did not find in vitro and in vivo scientific evaluation of F. assa-foetida L. and Ar. catechu extracts against H. contortus models.

In Pakistan, information related to medicinal plants used for the treatment of parasitic infections and other illnesses transfers from one generation to another. However, this indigenous knowledge varies greatly in different ethnic groups and from region to region due to local variation in the availability of plants (Hussain et al., 2008Hussain A, Khan MN, Iqbal Z, Sajid MS. An account of the botanical anthelmintics used in traditional veterinary practices in Sahiwal district of Punjab, Pakistan. J Ethnopharmacol 2008; 119(1): 185-190. http://dx.doi.org/10.1016/j.jep.2008.06.034. PMid:18657602.
http://dx.doi.org/10.1016/j.jep.2008.06....
). Due to geographic and climatic diversity in Pakistan, a wide variety of medicinal plants has been reported from different parts of the country (Iqbal et al., 2004Iqbal Z, Lateef M, Ashraf M, Jabbar A. Anthelmintic activity of Artemisia brevifolia in sheep. J Ethnopharmacol 2004; 93(2-3): 265-268. http://dx.doi.org/10.1016/j.jep.2004.03.046. PMid:15234763.
http://dx.doi.org/10.1016/j.jep.2004.03....
; Muhammad et al., 2005Muhammad G, Khan MZ, Hussain MH, Iqbal Z, Iqbal M, Athar M. Ethnoveterinary practices of owners of pneumatic-cart pulling camels in Faisalabad (Pakistan). J Ethnopharmacol 2005; 97(2): 241-246. http://dx.doi.org/10.1016/j.jep.2004.11.008. PMid:15707760.
http://dx.doi.org/10.1016/j.jep.2004.11....
; Hussain et al., 2008Hussain A, Khan MN, Iqbal Z, Sajid MS. An account of the botanical anthelmintics used in traditional veterinary practices in Sahiwal district of Punjab, Pakistan. J Ethnopharmacol 2008; 119(1): 185-190. http://dx.doi.org/10.1016/j.jep.2008.06.034. PMid:18657602.
http://dx.doi.org/10.1016/j.jep.2008.06....
; Goraya et al., 2013Goraya K, Iqbal Z, Sajid MS, Muhammad G, Ain QU, Saleem M. Diversity of flora used for the cure of equine diseases in selected peri-urban areas of Punjab, Pakistan. J Ethnobiol Ethnomed 2013; 9(1): 70. http://dx.doi.org/10.1186/1746-4269-9-70. PMid:24283263.
http://dx.doi.org/10.1186/1746-4269-9-70...
; Badar et al., 2017Badar N, Iqbal Z, Sajid MS, Rizwan HM, Jabbar A, Babar W, et al. Documentation of ethnoveterinary practices in district Jhang, Pakistan. J Anim Plant Sci 2017; 27(2): 398-406.). Most of these plants are used by native healers without specific knowledge about the active ingredients. It is important to document and validate plants used by local communities for medicinal purposes before their disappearance due to environmental and technological changes (Goraya et al., 2013Goraya K, Iqbal Z, Sajid MS, Muhammad G, Ain QU, Saleem M. Diversity of flora used for the cure of equine diseases in selected peri-urban areas of Punjab, Pakistan. J Ethnobiol Ethnomed 2013; 9(1): 70. http://dx.doi.org/10.1186/1746-4269-9-70. PMid:24283263.
http://dx.doi.org/10.1186/1746-4269-9-70...
; Badar et al., 2017Badar N, Iqbal Z, Sajid MS, Rizwan HM, Jabbar A, Babar W, et al. Documentation of ethnoveterinary practices in district Jhang, Pakistan. J Anim Plant Sci 2017; 27(2): 398-406.). In connection with earlier studies, the objective of this work was to determine the anthelmintic activities of the leaves of A. donax L., latex of F. assa-foetida L. and seeds of Ar. catechu L through in vitro and in vivo assays against H. contortus models.

Materials and Methods

Collection of plant materials

The plants A. donax L., Ar. catechu L. and F. assa-foetida L were selected during a documentation survey of the herbs used for the treatment of helminthiasis in animals (Badar et al., 2017Badar N, Iqbal Z, Sajid MS, Rizwan HM, Jabbar A, Babar W, et al. Documentation of ethnoveterinary practices in district Jhang, Pakistan. J Anim Plant Sci 2017; 27(2): 398-406.). Briefly, participatory epidemiological procedures were adopted to involve the local healers as respondents for the identification of the most commonly used plants for the treatment of worm infection in animals of the area (Hussain et al., 2008Hussain A, Khan MN, Iqbal Z, Sajid MS. An account of the botanical anthelmintics used in traditional veterinary practices in Sahiwal district of Punjab, Pakistan. J Ethnopharmacol 2008; 119(1): 185-190. http://dx.doi.org/10.1016/j.jep.2008.06.034. PMid:18657602.
http://dx.doi.org/10.1016/j.jep.2008.06....
). The leaves of A. donax L. (voucher No. Jg8/2006), latex of F. assa-foetida L. (voucher No. Jg24/2006), and seeds of Ar. catechu L. (voucher No. Jg7/2006) were collected from the market and field of district Jhang, Punjab, Pakistan. Identification of the plants was done by Prof. Dr. Abdul Wahid from the Department of Botany, University of Agriculture, Faisalabad, Pakistan.

Extraction and fractionation

Extraction

The leaves of A. donax L., latex of F. assa-foetida L. and seeds of Ar. catechu L. were dried at the room temperature. The dried specimens were ground to powder with an electric grinder. The powder was soaked for 3 days in an aqueous-methanol 30:70 suspension for the preparation of crude aqueous-methanol extracts (CAME) and filtered through muslin cloth and filter paper. The whole procedure was repeated thrice and then by the use of rotary evaporator at 40°C and low pressure, the combined filtrate was evaporated to get CAME (Tabassam et al., 2008Tabassam SM, Iqbal Z, Jabbar A, Sindhu ZD, Chattha AI. Efficacy of crude neem seed kernel extracts against natural infestation of Sarcoptes scabiei var. ovis. J Ethnopharmacol 2008; 115(2): 284-287. http://dx.doi.org/10.1016/j.jep.2007.10.003. PMid:18023309.
http://dx.doi.org/10.1016/j.jep.2007.10....
).

Fractionation

Organic solvents, including ethyl acetate, chloroform, and petroleum spirit, were used to fractionate the crude extracts as described (Williamson et al., 1998Williamson EM, Okpako DT, Evans FJ. Preparation of plant material. Pharmacological methods in phytotherapy research. Chichester, UK: John Wiley & Sons; 1998.). Briefly, after dissolving crude extract (20 g) in the distilled water (20 mL), and petroleum spirit (60 mL) in a funnel, was shaken vigorously. To separate the layers of petroleum spirit and the distilled water, the funnel was kept undisturbed for about 30 min. Another 60 mL petroleum spirit was added and the procedure was repeated until a clear petroleum spirit layer was obtained. All separated petroleum spirit layers were pooled and evaporated in a rotary evaporator. Fractions of chloroform and ethyl acetate were obtained by adopting the same procedure.

Parasitological procedures

Procurement of adult Haemonchus contortus

Following the methods of Alawa et al. (2003)Alawa CBI, Adamu AM, Gefu JO, Ajanusi OJ, Abdu PA, Chiezey NP, et al. In vitro screening of two Nigerian medicinal plants (Vernonia amygdalina and Annona senegalensis) for anthelmintic activity. Vet Parasitol 2003; 113(1): 73-81. http://dx.doi.org/10.1016/S0304-4017(03)00040-2. PMid:12651218.
http://dx.doi.org/10.1016/S0304-4017(03)...
, adult Haemonchus (H.) contortus worms were obtained from the abomasal contents of slaughtered sheep. Each abomasum was separated from the small intestine and shifted to the Ethnoveterinary Research Center at the Department of Parasitology, University of Agriculture, Faisalabad, Pakistan and examined within 3 h of slaughter. The abomasum was split open and their contents washed gently with water in separate buckets and collected. The abomasum was divided longitudinally and the whole mucosa was examined carefully. The worms were collected gently from the infected abomasum with the help of forceps. Some of the worms were separated to use in adult motility assay; whereas, from the remaining worms, females were separated and crushed using a pestle and mortar to liberate the eggs to be used in egg hatch assays.

Adult Motility Assay (AMA)

CAME and fractions of chloroform, ethyl acetate and petroleum spirit at concentrations of 50, 25, 12.5, 6.25, 3.12 and 1.56 mg mL-1 in phosphate-buffered saline (PBS) along with levamisole (0.55 mg mL-1; positive control) and PBS (negative control) were prepared. Adult live Haemonchus contortus were collected from abomasa of sheep procured from the local slaughter house. Minimum 10 worms were separated into individual Petri dishes and exposed to the above treatments. Each treatment was repeated thrice at the room temperature and the number of surviving and dead worms was observed at 0, 2, 4, 6, 8, 10, 12, and 24 hours post-treatment as described elsewhere (Lateef et al., 2003Lateef M, Iqbal Z, Khan MN, Akhtar MS, Jabbar A. Anthelmintic activity of Adhatoda vesica roots. Int J Agric Biol 2003; 5(1): 86-90.; Zaman et al., 2012Zaman MA, Iqbal Z, Khan MN, Muhammad G. Anthelmintic activity of a herbal formulation against gastrointestinal nematodes of sheep. Pak Vet J 2012; 32(1): 117-121.).

Egg Hatch Assay (EHA)

Eggs were separated from female H. contortus by triturating them in a pestle and mortar. About 250 eggs in 1.5 mL water were placed in each well of a 24-well plate. The CAME and fractions of chloroform, ethyl acetate and petroleum spirit along with albendazole (positive control) and PBS (negative control) were used at 1.2, 12, 120, 1200 and 12000 µg mL-1. The plate was incubated at 28°C for 36 hr. After incubation, eggs and larvae (dead or alive) were counted using an inverted microscope (Coles et al., 1992Coles GC, Bauer C, Borgsteede FH, Geerts S, Klei TR, Taylor MA, et al. World Association for the Advancement of Veterinary Parasitology (W.A.A.V.P) methods for the detection of anthelmintic resistance in nematodes of veterinary importance. Vet Parasitol 1992; 44(1-2): 35-44. http://dx.doi.org/10.1016/0304-4017(92)90141-U. PMid:1441190.
http://dx.doi.org/10.1016/0304-4017(92)9...
; Zaman et al., 2012Zaman MA, Iqbal Z, Khan MN, Muhammad G. Anthelmintic activity of a herbal formulation against gastrointestinal nematodes of sheep. Pak Vet J 2012; 32(1): 117-121.). Each treatment was repeated in triplicate.

Faecal Egg Count Reduction Test (FECRT)

Naturally H. contortus- infected sheep (4-8 months-old) were selected for FECRT. The study was conducted at a local farm of Faisalabad district, Punjab, Pakistan. Animals were screened through the “Modified McMaster test”. This study was approved by Research Ethics Committee, Faculty of Veterinary Science, University of Agriculture, Faisalabad, Pakistan. Standard guidelines for the institutional animal care and use (IACU), University of Agriculture, Faisalabad, Pakistan were followed. Sheep were divided into 20 groups of 5 animals each through a complete randomized design. Groups 1 and 2 were treated with PBS (negative control) and levamisole (positive control) at 7.5 mg kg-1 body weight, respectively. Groups 3-5 were treated with the crude powder (CP) of A. donax at 1, 4, and 7 g kg-1 body weight and groups 6-8 were treated with the CAME of A. donax at 1 g, 4 g, 7 g kg-1 body weight, respectively. Groups 9-11 were treated with the CP of F. assa-foetida at 0.33, 0.66, and 1 g kg-1 body weight, respectively and groups 12-14 were treated with the CAME of F. assa-foetida at 0.33, 0.66, and 1 g kg-1 body weight, respectively. Similarly, groups 15-17 were treated with the CP of Ar. catechu at 0.33, 0.66, and 1 g kg-1 body weight, respectively, and groups 18-20 were treated with the CAME of Ar. catechu at 0.33, 0.66, and 1 g kg-1 body weight, respectively. Each treatment was given once per os and faecal egg counts were done on days 0, 4, 8, and 12 post-treatment.

Statistical analyses

A characteristic sigmoid dose-response curve was transformed into a linear function (to get a constant or fixed rate of hatching with the increase concentration of different fractions) via the probit transformation of the egg hatch test (EHT) data. Probit analysis was used to calculate the lethal concentration 50 (LC50) of each extract for the EHT (Hubert & Kerboeuf, 1992Hubert J, Kerboeuf D. A microlarval development assay for the detection of anthelmintic resistance in sheep nematodes. Vet Rec 1992; 130(20): 442-446. http://dx.doi.org/10.1136/vr.130.20.442. PMid:1621342.
http://dx.doi.org/10.1136/vr.130.20.442...
). The Regression analysis was used to determine the effect of independent variables(concentration level) on the dependent variable (hatching) as described by Zaman et al. (2012)Zaman MA, Iqbal Z, Khan MN, Muhammad G. Anthelmintic activity of a herbal formulation against gastrointestinal nematodes of sheep. Pak Vet J 2012; 32(1): 117-121.. The adult motility assay (AMA) data were analyzed by the Duncan's multiple range (DMR) test to compare means of the number of dead worms. For FECRT, the results were expressed as eggs per gram (Mean±SEM) of faeces and means were compared by using DMR Test. All statistical analyses were carried out using SAS software (SAS, 1998Statistical Institute – SAS. Statistical analysis system: user’s guide. North Carolina, USA: Statistical Institute; 1998.).

Results

In vitro anthelmintic activity

In the AMA, 100% mortality (at 10th hr post-exposure) of H. contortus was noted with the CAME and ethyl acetate fractions of F. assa-foetida at 12.5-50 mg mL-1. The ethyl acetate fraction of A. donax showed maximum efficacy, followed in order by the fractions of chloroform, CAME and petroleum spirit. Among different fractions of Ar. Catechu, CAME showed the maximum mortality, followed in order by the fractions of ethyl acetate, chloroform and petroleum spirit. Levamisole caused 100% worm mortality within 2 hr post-exposure. The results of the AMA for the various treatments were observed as given in Table 1.

Table 1
Comparative efficacies of the crude aqueous methanolic extract and different fractions of Arundo donax (leaves), Ferula assa-foetida (latex) and Areca catechu (seeds) on the survival of Haemonchus contortus worms.

In the EHA, the CAME of F. assa-foetida showed the maximum ovicidal activity (LC50 =16.9 µg mL-1), followed in order by those of Ar. catechu and A. donax. The chloroform fractions of F. assa-foetida and A. donax showed more ovicidal activity than that of Ar. catechu. Both the regression values and their correlations regarding the effects of the treatments on egg hatching indicated that a better concentration-dependent effect was found with the CAME of F. assa-foetida compared to those of Ar. catechu and A. donax. The concentration-dependent effects of petroleum spirit fractions were observed as given in Figure 1.

Figure 1
In vitro hatching (%) of Haemonchus contortus eggs through crude aqueous methanol extract (CAME) and fractions of Ferula (F.) assa-foetida, Areca (Ar.) catechu and Arundo (A.) donax.

In vivo anthelmintic activity

Both the crude powder (CP) and crude aqueous methanol extracts (CAME) of the tested plants showed dose-dependent, variable levels of anthelmintic activity (Table 2). The anthelmintic efficacy of the CP of the plants ranged from 5.4% to 50.5%. The highest reduction (50.5%) in faecal egg count was recorded for A. donax at 7 g kg-1 than for F. assa-foetida at 1 g kg-1. The anthelmintic efficacies of the CAME preparations were not consistent with those recorded for the CP. The effects of all plant treatments (12 days post-treatment) were statistically different from untreated controls (P<0.05). Differences in eggs per gram (EPG) values of the levamisole-treated group from those of plant extract-treated groups were statistically significant (P<0.05). However, differences in EPG values of the groups treated with different doses of the plant extracts were not statistically significant (P>0.05) on days 8 and 12 post-treatment, treated with CP of F. assa-foetida and Ar. catechu.

Table 2
Faecal egg count reduction (Mean±SEM) in naturally infected sheep population with gastrointestinal parasites through crude powder and crude aqueous methanol extracts of the leaves of Arundo donax, latex of Ferula assa-foetida and seeds of Areca catechu in comparison with Levamisole

Discussion

Parasitic infection is a key problem responsible for reduction in the productivity of animals, partly due to poor feed utilization (Pedreira et al., 2006Pedreira J, Paz-Silva A, Sanchez-Andrade R, Suárez JL, Arias M, Lomba C, et al. Prevalence of gastrointestinal parasites in sheep and parasite-control practices in NW Spain. Prev Vet Med 2006; 75(1-2): 56-62. http://dx.doi.org/10.1016/j.prevetmed.2006.01.011. PMid:16488032.
http://dx.doi.org/10.1016/j.prevetmed.20...
). H. contortus is one of the principal parasitic nematodes of small ruminants, and the cause of huge economic losses (Radostits et al., 2000Radostits OM, Gay CC, Blood DC, Hinchcliff KW. Veterinary Medicine: a Textbook of the Diseases of Cattle, Sheep, Pigs, Goats and Horses. 9th ed. London, UK; W.B. Saunders Company Ltd:; 2000.). The control of haemonchosis and other parasites relies mainly on the use of anthelmintics (Khan et al., 2017Khan MN, Sajid MS, Rizwan HM, Qudoos A, Abbas RZ, Riaz M, et al. Comparative efficacy of six anthelmintic treatments against natural infection of Fasciola species in sheep. Pak Vet J 2017; 37(1): 65-68.). The efficacy of these chemicals is threatened by the development and spread of resistance. However, success in the development of new drugs for parasitic infections is limited (Kaminsky et al., 2008Kaminsky R, Ducray P, Jung M, Clover R, Rufener L, Bouvier J, et al. A new class of anthelmintics effective against drug-resistant nematodes. Nature 2008; 452(7184): 176-180. http://dx.doi.org/10.1038/nature06722. PMid:18337814.
http://dx.doi.org/10.1038/nature06722...
; Kumarasingha et al., 2016Kumarasingha R, Preston S, Yeo TC, Lim DSL, Tu CL, Palombo EA, et al. Anthelmintic activity of selected ethno-medicinal plant extracts on parasitic stages of Haemonchus contortus. Parasit Vectors 2016; 9(1): 187. http://dx.doi.org/10.1186/s13071-016-1458-9. PMid:27036205.
http://dx.doi.org/10.1186/s13071-016-145...
). Searching for new, safe, and effective anthelmintic compounds from plants is a promising way to control parasitic infection (Waller et al., 2001Waller PJ, Bernes G, Thamsborg SM, Sukura A, Richter SH, Ingebrigtsen K, et al. Plants as de-worming agents of livestock in the Nordic countries: historical perspective, popular beliefs and prospects for the future. Acta Vet Scand 2001; 42(1): 31-44. http://dx.doi.org/10.1186/1751-0147-42-31. PMid:11455900.
http://dx.doi.org/10.1186/1751-0147-42-3...
; Kumarasingha et al., 2016Kumarasingha R, Preston S, Yeo TC, Lim DSL, Tu CL, Palombo EA, et al. Anthelmintic activity of selected ethno-medicinal plant extracts on parasitic stages of Haemonchus contortus. Parasit Vectors 2016; 9(1): 187. http://dx.doi.org/10.1186/s13071-016-1458-9. PMid:27036205.
http://dx.doi.org/10.1186/s13071-016-145...
). Nature has provided a variety of medicinal plants, e.g., Trachyspermum ammi (L.) Sprague, Punica granatum L., Nicotiana tabacum L., Artemisia brevifolia, Allium sativum, Ferula asafoetida, Syzygium aromaticum and Withania coagulans Dunal, to cure parasitic diseases. The chemical compounds present in these plants have different solubilities in various solvents (Iqbal et al., 2004Iqbal Z, Lateef M, Ashraf M, Jabbar A. Anthelmintic activity of Artemisia brevifolia in sheep. J Ethnopharmacol 2004; 93(2-3): 265-268. http://dx.doi.org/10.1016/j.jep.2004.03.046. PMid:15234763.
http://dx.doi.org/10.1016/j.jep.2004.03....
; Kumar & Singh, 2014Kumar P, Singh DK. In vitro anthelmintic activity of Allium sativum, Ferula asafoetida, Syzygium aromaticum and their active components against Fasciola gigantica. J Biol Earth Sci 2014; 4(1): 57-65.; Atma et al., 2017Atma W, Larouci M, Meddah B, Benabdeli K, Sonnet P. Evaluation of the phytoremediation potential of Arundo donax L. for nickel-contaminated soil. Int J Phytoremediation 2017; 19(4): 377-386. http://dx.doi.org/10.1080/15226514.2016.1225291. PMid:27592714.
http://dx.doi.org/10.1080/15226514.2016....
). Different solvents can dissolve different chemical compounds (solutes) that underlie biological activities (Iqbal et al., 2007Iqbal Z, Sarwar M, Jabbar A, Ahmed S, Nisa M, Sajid MS, et al. Direct and indirect anthelmintic effects of condensed tannins in sheep. Vet Parasitol 2007; 144(1-2): 125-131. http://dx.doi.org/10.1016/j.vetpar.2006.09.035. PMid:17097807.
http://dx.doi.org/10.1016/j.vetpar.2006....
; McGaw et al., 2007McGaw LJ, Van der Merwe D, Eloff JN. In vitro anthelmintic, antibacterial and cytotoxic effects of extracts from plants used in South African ethnoveterinary medicine. Vet J 2007; 173(2): 366-372. http://dx.doi.org/10.1016/j.tvjl.2005.09.004. PMid:16239113.
http://dx.doi.org/10.1016/j.tvjl.2005.09...
; Altemimi et al., 2017Altemimi A, Lakhssassi N, Baharlouei A, Watson DG, Lightfoot DA. Phytochemicals: extraction, isolation, and identification of bioactive compounds from plant extracts. Plants 2017; 6(4): 42. http://dx.doi.org/10.3390/plants6040042. PMid:28937585.
http://dx.doi.org/10.3390/plants6040042...
). Such studies will help to identify potential candidates for alternative control through plant-based, novel anthelmintics. In the present study, solvents were used to fractionate plant extracts in order to identify those with the highest activity against the H. contortus model.

Various fractions of F. assa-foetida have been isolated, including gum, resins, volatile oils, coumarin derivatives and various monoterpenes, ferulic acid, farnesiferoles, disulfides, symmetric trisulfides and tetrasulfides (Hofer et al., 1984Hofer O, Widhalm M, Greger H. Circular dichroism of sesquiterpene-umbelliferone ethers and structure elucidation of a new derivative isolated from the gum resin ‘Asa Foetida’. Monatsh Chem 1984; 115(10): 1207-1218. http://dx.doi.org/10.1007/BF00809352.
http://dx.doi.org/10.1007/BF00809352...
; Kajimoto et al., 1989Kajimoto T, Yahiro K, Nohara T. Sesquiterpenoid and disulphide derivatives from Ferula assa-foetida. Phytochemistry 1989; 28(6): 1761-1763. http://dx.doi.org/10.1016/S0031-9422(00)97841-5.
http://dx.doi.org/10.1016/S0031-9422(00)...
; Kapoor, 1990Kapoor LD. Areca catechu Linn. Handbook of Ayurvedic Medicinal Plants. Boca Raton, Florida, USA: CRC Press; 1990.). The resin of F. assa-foetida has various properties, such as anticoagulant, smooth muscle relaxant, anti-diabetic, anticarcinogenic, antioxidant, antispasmodic, antihepatotoxic, antiulcerogenic, anticholesterolemic, anti-inflammatory, antifertility, antifungal, antiparasitic and anthelmintic (Kareparamban et al., 2012Kareparamban JA, Nikam PH, Jadhav AP, Kadam VJ. Ferula foetida “Hing”: a review. Res J Pharm Biol Chem Sci 2012; 3(2): 775-786., Amalraj & Gopi, 2016Amalraj A, Gopi S. Biological activities and medicinal properties of Asa foetida: a review. J Tradit Complement Med 2016; 7(3): 347-359. http://dx.doi.org/10.1016/j.jtcme.2016.11.004. PMid:28725631.
http://dx.doi.org/10.1016/j.jtcme.2016.1...
). The gum extract of F. assa-foetida has been used to cure diarrhea, constipation, abdominal pain, and parasitic infections (Fernch, 1971Fernch D. Ethnobothany of the Umbelliferae. In: Heywood VH. The chemistry and biology of the umberifella. London, UK: Academic Press; 1971. p. 285-412.). Inhibition of the growth of Shigella sonnei and Staphylococcus aureus has also been reported by the gum extract of F. assa-foetida (Kapoor, 1990Kapoor LD. Areca catechu Linn. Handbook of Ayurvedic Medicinal Plants. Boca Raton, Florida, USA: CRC Press; 1990.). Gundamaraju (2013)Gundamaraju R. Evaluation of anti-helmintic activity of Ferula foetida “Hing- A natural Indian spice” aqueous extract. Asian Pac J Trop Dis 2013; 3(3): 189-191. http://dx.doi.org/10.1016/S2222-1808(13)60038-9.
http://dx.doi.org/10.1016/S2222-1808(13)...
reported significant anthelmintic activity of F. assa-foetida at a concentration of 100 mg mL-1. Paralysis, as well as lethality, of an aqueous extract of F. assa-foetida was comparable to piperazine citrate at a concentration of 100 mg mL-1. Polyphenolic compounds and flavonoids are the main phytochemical components of crude extracts. Polyphenolic compounds such as tannins have been documented as anthelmintics (Bate-Smith, 1962Bate-Smith EC. The phenolic constituents of plants and their taxonomic significance. I. Dicotyledons. Bot J Linn Soc 1962; 58(371): 95-103. http://dx.doi.org/10.1111/j.1095-8339.1962.tb00890.x.
http://dx.doi.org/10.1111/j.1095-8339.19...
; Iqbal et al., 2007Iqbal Z, Sarwar M, Jabbar A, Ahmed S, Nisa M, Sajid MS, et al. Direct and indirect anthelmintic effects of condensed tannins in sheep. Vet Parasitol 2007; 144(1-2): 125-131. http://dx.doi.org/10.1016/j.vetpar.2006.09.035. PMid:17097807.
http://dx.doi.org/10.1016/j.vetpar.2006....
). Kumar & Singh (2014)Kumar P, Singh DK. In vitro anthelmintic activity of Allium sativum, Ferula asafoetida, Syzygium aromaticum and their active components against Fasciola gigantica. J Biol Earth Sci 2014; 4(1): 57-65. reported the LC50 of an ethanolic extract of the dried-root latex powder of F. assa-foetida (4.88 mg mL-1) followed in order by the dried clove powder of A. sativum (3.48 mg mL-1), and dried flower bud powder of Syzygium aromaticum (2.95 mg mL-1) against Fasciola gigantica. The possible anthelmintic effects of F. assa-foetida may be due to interference with energy generation in parasites by uncoupling oxidative phosphorylation or due to presence of tannins in the extracts that can bind to glycoprotein on the cuticle of the parasite and cause death (Mali & Wadekar, 2008Mali RG, Wadekar RR. In vitro anthelmintic activity of Baliospermum montanum Muell. Arg roots. Indian J Pharm Sci 2008; 70(1): 131-133. http://dx.doi.org/10.4103/0250-474X.40352. PMid:20390101.
http://dx.doi.org/10.4103/0250-474X.4035...
).

Areca catechu nuts were used as anthelmintics and appear in the British pharmacopoeia for the treatment of parasitic infections (Raghavan & Baruah, 1958Raghavan V, Baruah HK. Arecanut: India’s popular masticatory - history, chemistry and utilization. Econ Bot 1958; 12(4): 315-345. http://dx.doi.org/10.1007/BF02860022.
http://dx.doi.org/10.1007/BF02860022...
). The nuts of Ar. catechu contain tannins, gallic acid, oil gum, volatile oils, lignins (Huang, 1992Huang KC. Anthelmintics. In: The Pharmacology of Chinese Herbs. Boca Raton, Florida, USA: CRC press; 1992. p. 321-333.), alkaloids, phenolic compounds, beta-sitosterol, catechins, amino acids and various saline substances (Duke, 1992Duke JA. Handbook of phytochemical constituents of GRAS herbs and other economic plants. London, UK: CRC Press; 1992.; Wang et al., 1997Wang CK, Lee WH, Peng CH. Contents of phenolics and alkaloids in Areca catechu Linn. during maturation. J Agric Food Chem 1997; 45(4): 1185-1188. http://dx.doi.org/10.1021/jf960547q.
http://dx.doi.org/10.1021/jf960547q...
; Chu, 2001Chu NS. Effects of betel chewing on the central and autonomic nervous systems. J Biomed Sci 2001; 8(3): 229-236. http://dx.doi.org/10.1007/BF02256596. PMid:11385294.
http://dx.doi.org/10.1007/BF02256596...
). An ethanolic extract of Ar. catechu significantly inhibited fumarate reductase and succinate dehydrogenase activities in Cotylophoron cotylophorum (Dhanraj & Veerakumari, 2016Dhanraj KM, Veerakumari L. Effect of ethanol extract of Areca catechu on fumarate reductase and succinate dehydrogenase of Cotylophoron cotylophorum. Int J Res Develop Pharm Development Life Sci 2016;5(3): 2117-2123.). The combined efficacy of Ar. catechu and Anredera (An.) cordifolia showed significant elimination of Ascaridia galli (60%) in chickens and reduced EPG of faeces. However, in a group treated with alkaloids and saponins contained in Ar. catechu and An. cordifolia showed antagonistic activity (Prastowo et al., 2017Prastowo J, Herawati O, Ariyadi B, Kurniasih. Effects of Areca catechu seed and Anredera cordifolia leaf on Ascaridia galli infection in the domestic chicken (Gallus gallus domesticus). Int J Poult Sci 2017; 16(12): 494-499. http://dx.doi.org/10.3923/ijps.2017.494.499.
http://dx.doi.org/10.3923/ijps.2017.494....
). Baby & Raphael (2014)Baby AA, Raphael RG. Potential antimicrobial, anthelmintic and antioxidant properties of Areca catechu L. root. Int J Pharm Pharm Sci 2014; 6(6): 486-489. reported dose-dependent anthelmintic activity of ethanolic extract of Ar. catechu root against Pheretima posthuma. The results of the study conducted by Yamson et al. (2019)Yamson EC, Tubalinal GASP, Viloria VV, Mingala CN. Anthelmintic effect of betel nut (Areca catechu) and neem (Azadirachta indica) extract against liver fluke (Fasciola spp.). J Adv Vet Anim Res 2019; 6(1): 44-49. http://dx.doi.org/10.5455/javar.2019.e310. PMid:31453169.
http://dx.doi.org/10.5455/javar.2019.e31...
revealed that there was no movement of Fasciola sp. when exposed to 40% ethanol extract Ar. catechu. Most of the pharmacological and biological effects, such as inhibition of acetylcholinesterase, stimulation of the relaxed bowel and cholinomimetic effects of Ar. catechu, are due to the condensed tannins (Kapoor, 1990Kapoor LD. Areca catechu Linn. Handbook of Ayurvedic Medicinal Plants. Boca Raton, Florida, USA: CRC Press; 1990.; Amudhan et al., 2012Amudhan MS, Begum VH, Hebbar KB. A review on phytochemical and pharmacological potential of Areca catechu L. seed. Int J Pharm Sci Res 2012; 3: 4151-4157.). Direct and/or indirect pharmacological effects of the condensed tannins on the GI parasites have been reported (i.e., Butter et al., 2000Butter NL, Dawson JM, Wakelin D, Buttery PJ. Effect of dietary tannin and protein concentration on nematode infection (Trichostrongylus colubriformis) in lambs. J Agric Sci 2000; 134(1): 89-99. http://dx.doi.org/10.1017/S0021859699007315.
http://dx.doi.org/10.1017/S0021859699007...
; Molan et al., 2000Molan AL, Waghorn GC, Min BR, McNabb WC. The effect of condensed tannins from seven herbages on Trichostrongylus colubriformis larval migration in vitro. Folia Parasitol (Praha) 2000; 47(1): 39-44. http://dx.doi.org/10.14411/fp.2000.007. PMid:10833014.
http://dx.doi.org/10.14411/fp.2000.007...
; Iqbal et al., 2007Iqbal Z, Sarwar M, Jabbar A, Ahmed S, Nisa M, Sajid MS, et al. Direct and indirect anthelmintic effects of condensed tannins in sheep. Vet Parasitol 2007; 144(1-2): 125-131. http://dx.doi.org/10.1016/j.vetpar.2006.09.035. PMid:17097807.
http://dx.doi.org/10.1016/j.vetpar.2006....
; Naumann et al., 2017Naumann HD, Tedeschi LO, Zeller WE, Huntley NF. The role of condensed tannins in ruminant animal production: advances, limitations and future directions. Rev Bras Zootec 2017; 46(12): 929-949. http://dx.doi.org/10.1590/s1806-92902017001200009.
http://dx.doi.org/10.1590/s1806-92902017...
). Condensed tannins can interfere with physiological functioning of parasites, development of infective larval stages and hatching of parasite eggs (Molan et al., 2000Molan AL, Waghorn GC, Min BR, McNabb WC. The effect of condensed tannins from seven herbages on Trichostrongylus colubriformis larval migration in vitro. Folia Parasitol (Praha) 2000; 47(1): 39-44. http://dx.doi.org/10.14411/fp.2000.007. PMid:10833014.
http://dx.doi.org/10.14411/fp.2000.007...
).

In Ayurvedic medicine, a decoction of A. donax was used as haemostatic, in toothache, as a diuretic and as an emollient (Passalacqua et al., 2007Passalacqua NG, Guarrera PM, De Fine G. Contribution to the knowledge of the folk plant medicine in Calabria region (Southern Italy). Fitoterapia 2007; 78(1): 52-68. http://dx.doi.org/10.1016/j.fitote.2006.07.005. PMid:17084993.
http://dx.doi.org/10.1016/j.fitote.2006....
). It is also used to diminish the secretion of milk and to stimulate menstrual discharge (Chopra et al., 1956Chopra RN, Nayyar SL, Chopra IC. Glossary of Indian medicinal plants. New Delhi, India: Council of Scientific and Industrial Research; 1956.). It is a potential biomass plant with an invasive potential in riparian areas (Nackley & Kim, 2015Nackley LL, Kim SH. A salt on the bioenergy and biological invasions debate: salinity tolerance of the invasive biomass feedstock Arundo donax. Glob Change Biol Bioenergy 2015; 7(4): 752-762. http://dx.doi.org/10.1111/gcbb.12184.
http://dx.doi.org/10.1111/gcbb.12184...
; Al-Snafi, 2015Al-Snafi AE. The constituents and biological effects of Arundo donax- a review. Int J Phytopharm Res 2015; 6(1): 34-40.) and is also used for phytoremediation in Ni-contaminated soils (Atma et al., 2017Atma W, Larouci M, Meddah B, Benabdeli K, Sonnet P. Evaluation of the phytoremediation potential of Arundo donax L. for nickel-contaminated soil. Int J Phytoremediation 2017; 19(4): 377-386. http://dx.doi.org/10.1080/15226514.2016.1225291. PMid:27592714.
http://dx.doi.org/10.1080/15226514.2016....
). Five indole-3-alkylamine bases (5-methoxy-N-methyl-tryptamine, bufotenidine, bufotenine, N-dimethyltryptamine, and dehydrobufotenine) have been isolated from rhizomes of A. donax (Ghosal et al., 1969Ghosal S, Dutta SK, Sanyal AK, Bhattacharya SK. Arundo donax L. (Grarninae): phytochemical and pharmacological evaluation. J Med Chem 1969; 12(3): 480-483. http://dx.doi.org/10.1021/jm00303a032. PMid:5815188.
http://dx.doi.org/10.1021/jm00303a032...
). Antispasmodic and hypotensive effects against serotonin, acetylcholine, and histamine-induced spasms by a defatted, ethanolic extract of A. donax rhizomes have been reported (Al-Snafi, 2018Al-Snafi AE. Medicinal plants affected contractility of smooth muscles- a review. IOSR J Pharm 2018; 8(11): 22-35.). In a study conducted by Sharatkumar et al. (2004)Sharatkumar S, Dhanachand C, Mohilal N. Study on the efficacy of certain medicinal plants on gastrointestinal helminths of cattle. Indian Vet J 2004; 81(5): 497-498., 25-50 mg mL-1 A. donax showed significant anthelmintic effects. The highest mortality of H. contortus after levamisole (used as a reference drug) was 56.7%, with the CAME of A. donax at the concentration of 50 mg mL-1 at 10 hr post-exposure. Among various fractions of A. donax against H. contortus, ethyl acetate was the most active, followed in order by chloroform, aqueous and petroleum spirits extracts. In a FECRT, 50.5% reduction of eggs was recorded by the CP of A. donax in naturally infected sheep. However, A. donax exhibited ovicidal activity, with LC50 of 200.1 μg mL-1. Around 55% efficacy of A. donax extracts was recorded against various GI parasites of cattle (Miles et al., 1993Miles DH, Tunsuwan K, Chittawong V, Kokpol U, Choudhary MI, Clardy J. Boll weevil antifeedants from Arundo donax. Phytochemistry 1993; 34(5): 1277-1279. http://dx.doi.org/10.1016/0031-9422(91)80015-S.
http://dx.doi.org/10.1016/0031-9422(91)8...
).

In conclusions, the preparations of F. assa-foetida were toxic to H. contortus in vitro. In the AMA and EHA, CAME showed maximum mortality and ovicidal activity. However, the ethyl acetate fraction of A. donax showed maximum efficacy against adult worms, while chloroform fractions of F. assa-foetida and A. donax showed maximum ovicidal activity. In vivo efficacy of CP and CAME in this study ranged from 5.4% to 50.5%. The higher reduction (50.5%) in faecal egg count was recorded for A. donax at the dose rate of 7 g kg-1. Differences in the values of EPG between the levamisole-treated group and those treated with plant extracts were statistically significant (P<0.05).

Acknowledgements

The study was partially-funded by the project titled “Documentation and evaluation of some traditionally used ethnobotanicals for their anthelmintic activity” No. 20-362/R&D/05, The Higher Education Commission, Islamabad, Pakistan. The authors thank the local farming community and personnel of the Livestock and Dairy Development Department of the selected sites for providing valuable information about commonly used plants against gastrointestinal parasitism and assistance in collection of the plants. Identification of the plants was done by Prof. Dr. Abdul Wahid from the Department of Botany, University of Agriculture, Faisalabad, Pakistan. Authors are very thankful to Emeritus Professor Dr. Timothy G. Geary, McGill University, McDonald Campus, Quebec, Canada, for English language revision of the manuscript.

  • How to cite: Badar SN, Iqbal Z, Sajid MS, Rizwan HM, Shareef M, Malik MA, et al. Comparative anthelmintic efficacy of Arundo donax, Areca catechu, and Ferula assa-foetida against Haemonchus contortus. Braz J Vet Parasitol 2021; 30(2): e001221. https://doi.org/10.1590/S1984-29612021028

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Publication Dates

  • Publication in this collection
    28 May 2021
  • Date of issue
    2021

History

  • Received
    18 Jan 2021
  • Accepted
    10 Mar 2021
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