Efficacy of Closantel, Ferula Asafetida and Common Dewormer Against Haemoncus Contortus in Small Ruminants

doi:10.21203/rs.3.rs-575840/v1

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Research Article

Efficacy of Closantel, Ferula Asafetida and Common Dewormer Against Haemoncus Contortus in Small Ruminants

https://doi.org/10.21203/rs.3.rs-575840/v1

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Background

Haemonchosis caused by H. contortus has been recognized as one of the main limitations to the beneficial goat farming, worldwide. Albendazole, Oxfendazole, and Ivermectin are the most common drugs used against Haemonchosis, development of allopathic drugs resistance with time, availability, and price are another critical issues; these are absent in herbal drugs. It is important to establish a most effective protocol for the treatment of Haemonchosis. Closantel and herbal plant Ferula asafetida are reported to be effective against roundworms infestation.

Methodology:

The current study was conducted on goat and sheep farms of Tehsil Pattoki District Kasur, Pakistan. Coprological examination was performed and animals n = 720 (n = 360 caprines & n = 360 Ovines) positive Haemonchosis were selected, each breed animals were divided into six groups (A-F), n = 60 animals were assigned in each group for therapeutic trails. By oral route, Albendazole, Oxfendazole, Ivermectin, Closantel and Ferula asafetida were given to A-E groups while F served as control. All animals treated with single dose and EPG performed by McMaster technique at day 0 (pre-treatment), 7th, and 14th (post-treatment).

Results

Closantel and Ferula asafetida showed the highest efficacy in both caprines and ovines.

Conclusion

Allopathic drug Closantel and herbal drug Ferula asafetida proved as an efficient dewormer against Haemonchosis.

Animal Science

Haemonchosis

Small ruminants

Resistance

Closantel

Ferula asafoetida

Haemonchosis in small ruminants is caused by blood sucking gastrointestinal roundworm, Haemonchus contortus (H. contortus), Adult parasite suck blood (0.05 ml blood/day/parasite) and damage intestinal walls of host, severe infestations lead to anaemia, hypo proteinemia, edema, pregnancy toxemia, diarrhea, severe emaciation and high mortality (Besier et al. 2016, Dey et al. 2019, Dutta et al. 2017). The prevalence of this parasite reported worldwide and in Pakistan prevalence range is 24.6%to 80.64%, that varies with season (Al-Shaibani et al. 2008, Asif et al. 2008, Dey, Zhang, Begum, Alim, Hu and Alam 2019, Qasim et al. 2016). Haemonchosis is one of the main limiting factors in beneficial small ruminants farming and Pakistan is standing at 3rd in production of goat population in the world (Shah et al. 2017). In Pakistan, losses are in billions in form of growth, milk, death in young animals, hair and wool production (Rizwan et al. 2017). Anthelmintic resistance (AR) is described as low efficacy of an anthelmintic against parasites population that is generally susceptible to that drug (Fleming et al. 2006). Among many species of gastrointestinal nematodes, resistance of variable degree has been developed due to use of frequent and low dose of same drug, and usually resistance appears with in ten years of introduction of new drug. Field populations of H. contortus species show resistance to major classes of anthelmintic like benzimidazole, macrocyclic lactons and imidazothiazoles. In Pakistancommonly used anthelmicts are albenzaloes and ivermectin, and anthelmintic resistance is the main challenge to treat Haemonchosis in animals (Iqbal et al. 2017, Kotze and Prichard 2016, Shalaby 2013).

Albendazole and Oxfendazole belongs to benzimidazole group of anthelmentics, and Ivermectin from Tetrahydropyramidine, showed high efficacy against nematodes in goats both in vivo and vitro trials but these drugs are used in huge way that resistance have been developed against them (Adediran and Uwalaka 2015, Das et al. 2015, Gonzalez et al. 2019, Kaplan and Vidyashankar 2012, Wang et al. 2017). Closantel is another drug reported very effective against H. contortus, and is preferred routine wise over benzimidazoles (Dixit et al. 2019, Westers et al. 2016). Resistance with allopathic is always an issue however, disadvantage of allopathic drugs is resistance that develop with time furthermore, availability and price is another issue whereas these issues are absent in case of herbal drugs. Many herbal drugs are effective against gastrointestinal nematodes (Hamad 2014).

Ferula asafoetida (Hing) is a herbal drug, it has traditionally been used for its anthelminthic properties in many countries such as Iran, China and Nepal, where it has been used for treating infection with intestinal parasites (Bhatnager et al. 2015, Şahin et al. 2003). This drug has not yet been tried against H. contortus, furthermore, this drug is easily available in Pakistan. In this study, we planned to document in vivo anthelmintic effect of Albendazole, Oxfendazole, Ivermectin, Closantel and crude aqueous extract of herbal drug Ferula asafoetida on H. contortus, gastrointestinal nematodes of Goats and Sheep.

2.1. Ethical approval and informed consent

Ethical approval for this study was otained from University of Veterinary and Animal Sciences (UVAS), Lahore, Pakistan.

2.2. Sampling area

The current study was conducted at the sheep and goats farms in Tehsil Pattoki, District Kasur, Punjab, Pakistan. It lies between 168 meters above the sea level; it is bordered by the Ravi River in the north-west and river Sutlej in the south-east. In district Kasur, mean temperature in summer is 45ºC, whereas in winter, it becomes as lower as 6ºC. The average rainfall in Kasur district ranges from 161.9 with an average of 0 mm.

2.3. Sample collection

Fecal Samples were collected from the animals which was not dewormed in the last 4 months showing signs of anemia, edema, diarrhea, loss of hairs and loss of weight during July to December 2018 according to Soulsby protocol which is considered as standard. Approximately 5 g fresh sample of faeces directly collected from the rectum of animals in a labelled plastic tube having ratio of 3:1 formalin (1 part of sample and 3 parts formalin). Gender and age were recorded for each animal and transported to University Diagnostic Lab, University of Veterinary and Animal Sciences Ravi campus Pattoki. The samples were kept in refrigerator until examination within 24 hours. These fecal samples were characterized into three scores, one for normal, two for semisolid and three for diarrheic feces (Rizwan, Sajid, Iqbal and Saqib 2017).

2.4. Grouping of animals

Total haemonchosis positive animals n = 720 (n = 360 Caprines & n = 360 Ovines) were there. On the number of Egg per Gram animals were divided into three different groups’ highly infected, moderate infected and light infected animals with H. contortus, their EPG was respectively more than 1200, 800 to1200 and 50 to 799 (8). For therapeutic trail animals were arranged into six groups (A-F) each containing 60 animals A-E groups were in therapeutic trail while F group was control group.

2.5. Fecal Egg Count (FEC)

Samples were preceded by using techniques of parasitology (sedimentation and flotation) and identified as positive samples. For quantitative examination to know egg’s concentration “McMaster technique” was done. By the identification of ova or egg of H. contortus were known. Egg of H. contortus is shown in Fig. 1.

2.6. Therapeutic trails

Group A, B, C, D, E were treated orally with albendazole, oxfendazole, Ivermectin, Closantel and Ferula asafetida by using the dose rate of 10 mg/ kg, 5mg/kg, 0.02 mg/kg, 10 mg/kg and 100 mg/kg respectively, while F was not treated with any medicine and kept as control. The animals of respective groups were treated at day 0 after sample collection. Fecal samples were collected at day 0 (pre- treatment), day 7 and at 14 (post treatment) of all animals (Dixit, Das, Dixit and Sharma 2019, dos Santos et al. 2017, Puspitasari et al. 2016). Control group was dewormed using Closantel on day 14th after sample collection. Ferula asafoetida plant was collected in July 2017, the plant was shade dried for several days. Whole plants were dried and ground in a hammer mill then soaked in water for seven days at room temp and dried powered was given at rate of 100mg/kg oral, procedure followed as (Tavassoli et al. 2018). Allocation of groups and treatments with their dose rates is given in the Table. I

2.7. Statistical analysis

The collected data set was imported into SPSS (SPSS Inc., Chicago, Illinois, USA) version 26.0. The Chi-Square (X²) was used for the efficacy while repeated measure ANOVA was used for the EPG count.

In the current study, the effectiveness of Albendazole, Oxfendazole, Ivermectin, Closantel, and Ferula asafoetida were evaluated against H. contortus on the basis of the decrease of EPG in caprine and ovine. The results showed that there was significantly higher efficacy of closantel (100%) as compared to other treatment drugs in caprine (Fig. 2). The data of drugs efficacy showed that in ovine, closantel, and ferula asafetida (99 & 87%; respectively) were highly effective as compared to other treatment and control group (Fig. 3).

Moreover, both in caprine and ovine, there was no significance difference of drugs efficacy between male and female in all treatment groups (Table 2). Similarly, there was no significance difference in the efficacy of drugs between different age groups in both caprine and ovine (Table 3).

Table 1

Experimental groups, drugs and route of administration (dose and route)
Group	Formulation	Trade Name	Company	Route	Dose rate
A	Albendazole	Valbazine	Pfizer	Per oral	10mg/ kg
B	Oxfendazole	Systamax	ICI	Per oral	5mg/kg
C	Ivermectin	Ivoteck drench	Star	Per oral	0.02mg/kg
D	Closantel	Closancare	advacare	Per oral	10mg/kg
E	Ferula asafetida	---	---	Per oral	100mg/kg
F	Control Group	---	---	---	---

Table 2

Drug efficacy against *Haemonchus contartus* in caprine and ovine with relationship between gender.
Treatment Groups	Caprine		Ovine
Treatment Groups	Male	Female	Male	Female
Group A	40	39	9	9
Group B	33	33	4	4
Group C	22	22	17	17
Group D	100	100	99	99
Group E	70	70	87	87
Group F	0	0	0	0
Group A to F show Albendazole, Oxfendazole, Ivermectin, Closantel, Ferula asafetida and control groups.

Table 3

Drug efficacy against *Haemonchus contartus* in caprine and ovine relationship between different age
Treatment groups	Caprine			Ovine
Treatment groups	0–2 Yrs	2–3 Yrs	3–4 Yrs	0–2 Yrs	2–3 Yrs	3–4 Yrs
Group A	39	39	39	8.5	9	9
Group B	33	32	33	4.3	4	4
Group C	23	22	22	18	17	17
Group D	100	100	100	99	99	99
Group E	70	70	70	87	87	87
Group F	0	0	0	0	0	0
Group A to F show Albendazole, Oxfendazole, Ivermectin, Closantel, Ferula asafetida and control groups. Drug efficacy against Haemonchus contratus in caprine and ovine show in percentage (%)

3.1. Effect of drugs on reduction of EPG on various days in caprine and ovine

The results of reduction in the EPG in caprine showed that there was significantly higher reduction of EPG in closantel treatment at days 7 and 14 (104.13 ± 28.11 & 0, respectively) compared with other treatment drugs. The lowest reduction of EPG was noticed in ivermectin group at days 7 and 14 (1257.00 ± 409.24 and 1096.50 ± 356.73, respectively) among all treatment drugs. Moreover in all treatment, groups of therapeutic trail showed a significant reduction of EPG from day 0 through day 14 as compared to control (F), as presented in (Table 4).

Table 4

Effect of drugs on reduction of EPG on various day in Caprine.
Treatment groups	Day 0	Day 7	Day 14
Group A	1215.12 ± 344.37^Z	969.88 ± 278.81^bcY	730.88 ± 203.06^bcX
Group B	1462.12 ± 481.06^Z	1164.63 ± 384.18^cdY	937.13 ± 322.68^cdX
Group C	1409.87 ± 456.21^Z	1257.00 ± 409.24^cdY	1096.50 ± 356.73^cdX
Group D	1508.75 ± 407.11^Z	104.13 ± 28.11^aY	0.00^aX
Group E	1468.00 ± 342.53^Z	498.50 ± 116.69^abY	439.13 ± 103.63^bX
Control	1466.75 ± 340.27	1465.63 ± 337.85^d	1537.13 ± 365.13^d
Group A to F show Albendazole, Oxfendazole, Ivermectin, Closantel, Ferula asafetida and control groups. a, b, c and d show the level of significance between treatment groups, while X, Y and Z show level of significance difference within treatment groups.

The result of reduction in the EPG in ovine showed that there was significantly higher reduction of EPG from day 0 to days 14 in closantel and ferula (1873.51 ± 638.91 to 17.75 ± 5.65 and 1873.63 ± 607.19 to 243.54 ± 78.91, respectively) treatment groups among treatment and control group. There was non-significant difference between Albendazole, Oxfendazole, and Ivermectin treatment in EPG reduction at days 7 and 14 (Table 5).

Table 5

Effect of drugs on reduction of EPG on various days in ovine.
Treatment groups	Day 0	Day 7	Day 14
Group A	1747.13 ± 679.41^Y	1663.62 ± 646.91^bY	1582.25 ± 632.73^bY
Group B	1816 ± 729.61^Y	1781 ± 716.18^bY	1734.75 ± 699.73^bY
Group C	1738.25 ± 632.46^Y	1573.81 ± 578.02^bY	1432.75 ± 466.61^bY
Group D	1873.51 ± 638.91^Y	147.13 ± 51.13^aX	17.75 ± 5.65^aX
Group E	1873.63 ± 607.19^Y	399.12 ± 143.97^aX	243.54 ± 78.91^aX
Control	1884.88 ± 622.89	1910 ± 607.49^b	1951.51 ± 630.06^b
Group A to F show Albendazole, Oxfendazole, Ivermectin, Closantel, Ferula asafetida and control groups. a and b show the level of significance between treatment groups, while X and Y show level of significance difference within treatment groups.

However, there was no significant difference in reduction of EPG between male and female caprine and ovine in all treatment groups (Figs. 4 and 5). Similarly, there was no significant difference in EPG reduction between 0–2, 2–3 and 3–4 years caprine and ovine in all treatment groups (Figs. 6 and 7).

Traditionally anthelmintics are considered as major part of the strategies for the control of helminth in grazing animals for the prevention of losses of production. Use of same, unselective and having similar mode of action anthelmintics for many years cause resistance in animals (Coles and Roush 1992, Islam et al. 2018). Therefore in this study four common and one herbal anthelmintics drugs were used to evaluate the efficacy and anthelmintics resistance in caprine and ovine. To the best of our knowledge this was the first study in which efficacy of anthelminics were determined in goats and sheep. The main finding of this study was (a) the overall efficacy of closantal was higher in caprine, however in ovine closantel and ferula asafetida had higher drug efficacy irrespective of age and gender. (b) The EPG reduction was higher in closantel in both caprine and ovine, while ferula asafetida significantly reduced EPG in ovine than caprine.

The result of the current study showed that in caprine the EPG reduction was highest at 7th and 14th days post treatment in colantal than other anthelmintics, therefore the efficacy of closantal was 100% than other anthelmintics against h. contortus. Our result had in agreement with previous result that closantal had higher efficacy and low anthelminitics resistant in goats (Das, Dixit, Nath, Agrawal and Dongre 2015, Uppal et al. 1993) who also found Closantel to be 100 % effective against Haemonchus contortus in goat by using 10mg/kg body weight of medicine. The (Dixit, Das, Dixit and Sharma 2019) found that closantel is 95.64% effective against H. contortus.

The reason of higher efficacy of closantal against H. contartus might be high bounding properties to blood protein that is important for oxidative phosphorylation in parasites, which decreased the adenosine triphosphate and nicotinamide in the mitochondria, therefore availablily of energy lower for parasites (Lanusse et al. 2009). However some previous studies observed that anthelminitics developed resistant against closantal (Baihaqi et al. 2019, Chandrawathani et al. 2013, Premaalatha et al. 2014). The result of current study showed that the efficacy of ferula asafetida was 70% that is lowered than closantal and higher than other anthelminitics. (Singh et al. 2016) reported that 100% efficacy of aqueous extract of Zanthoxylum armatum DC. Seeds against H. contartus. (Akhtar et al. 2000) observed that different medicinal plants had high efficacy against anthelminitcs in goats of Indo-Pakistan. The high efficacy of these medicinal plants might be the high contents of phenolic flavonoid and tannin that is improved the immune defense system of host to destroy the parasites (Atoui et al. 2005, Tiwary et al. 2007). The other reason of low anthelminitcs resistant is the high concentration of cooper and zinic in these plants, which is high wormicidal and indirect role in improving immune defence of host (Burke et al. 2005, Chartier et al. 2001, Waruiru et al. 2004). We found high anthelminitics resistant against albendazol, oxfendazol and ivermectin, that is in agreements with previous reports (Kumar Jaiswal et al. 2013, Maharshi et al. 2011, Uppal, Yadav and Bhushan 1993).

The finding of current study showed that closantal and ferula asafetida had higher efficacy against H. contortus in sheep. Our result in agreements with previous studies that closantal had high efficacy against H. contortus and other strongylieds in sheep (Westers, Jones-Bitton, Menzies, Van Leeuwen, Poljak and Peregrine 2016) and merino sheep (Kadam et al. 2009, Shahana 2013, Tramboo et al. 2017). observed 100% and 98% respectively efficacy of closantal against H. contortus at 8th and 14th days of post treatment in Indian sheep. The higher efficacy of closantal against H. contortus may be due to lowered population of H. contortus eggs and improvement of immune defense of sheep. In literature ivermectin had higher efficacy (99%) against strongly and H. contortus in sheep, when injected subcutaneously (Canga et al. 2009, Westers, Jones-Bitton, Menzies, Van Leeuwen, Poljak and Peregrine 2016). The reason of higher efficacy of subcutaneous ivermectin might be higher bioavailability of ivermectin to plasma. The recent review of literature reports that aqueous extract of herbal plants had higher efficacy against strongly And H. contortus (de Souza Chagas and da Silva Vieira 2007, Dongre et al. 2015).

The reason of higher efficacy of herbal plants due to increased quantity of cooper and zinic that is lethal for the growth and production worm eggs (Chartier, Soubirac, Pors, Silvestre, Hubert, Couquet and Cabaret 2001, Waruiru, Mutunu and Otieno 2004). Similar finding have reported by (41,50) that suppimentation of copper and zinic had significant reduction of strongly eggs in host. In the current study albendazol, oxfendazol and ivermectin had lowered efficacy. Our finding has in agreement with previous reports that lowered efficacy of albendaol and fenbendazol due to higher anthelminitics resistance in small ruminants (Baihaqi, Widiyono and Nurcahyo 2019, Shahardar et al. 2014, Westers, Jones-Bitton, Menzies, Van Leeuwen, Poljak and Peregrine 2016). The higher anthelminitics resistance was due to higher frequency of deworming and use of insufficient dose of anthelminitics (Sissay et al. 2006).

The finding of current study revealed that no significant difference in age and gender wise efficacy of anthelminitics in both goats and sheep. This might be due to similar improvements of immune defense system against H. contortus in host. The reducation of eggs of H. contortus was significantly higher in closantal and ferula asafetida in both goats and sheep regardless of age and gender. Our finding support the previous findings that higher reduction of eggs in closantal and plants extract and leaves (Dixit, Das, Dixit and Sharma 2019, Nabukenya et al. 2014, Tramboo, Shahardar, Allaie, Wani and Abbas 2017). Albendazol, oxfendazol and oraly ivermectin had lowered reduction of eggs in the current study. These might be due to low availability of drug in the plasma that might be more expression of resistan genes (Coles et al. 2006, Kenyon and Jackson 2012, Van den Brom et al. 2015). A longer term study would be necessary to fully evaluate the resistance genes in the H. contortus.

Finally in conclusion, the higher reduction of H. contortus eggs after the treatment of closantal in goats and in sheep, the higher reduction of H. contortus eggs after the treatment of closantal and ferula asafetida. Therefore, higher efficacy of closantal against H. contortus is displayed in both goats and sheep, however, ferula asafetida displayd only higher efficacy in sheep.

Ethics approval was taken from University of Veterinary and Animal Sciences Lahore

Ethical considerations

Ethical issues (including plagiarism, informed consent, misconduct, data fabrication and/or falsification, double publication and/or submission, redundancy, etc.) have been completely observed by the authors

Author contributions

HMI designed the project. Sampling, data collection, processing and interpretation of results were made by AJ, MZI the data analysis was made by MA and MNW. The manuscript was written by AJ, HS, MI and MSZG. All the authors read the manuscript and approved the contents.

Acknowledgement

The authors highly acknowledged the Central Laboratory complex (CLC) UVAS Ravi Campus Pattoki for providing research facilities to carry out some part of this work.

Conflict of interest

The authors declare no conflict of interest

Acknowledgment

We would like to express our full acknowledgement to all authors who contributed in this work

Author Contribution

All authors contributed equally

Funding

There was no source of funding for this study

Availability of data and materials

None.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests

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Editorial decision: Major revision with re-assessment
17 Aug, 2021
Reviewers invited by journal
29 Jun, 2021
Editor assigned by journal
01 Jun, 2021
First submitted to journal
29 May, 2021

You are reading this latest preprint version

Efficacy of Closantel, Ferula Asafetida and Common Dewormer Against Haemoncus Contortus in Small Ruminants

Status:

Version 1

Abstract

Background

Methodology:

Results

Conclusion

Figures

1. Introduction

2. Material And Method

2.1. Ethical approval and informed consent

2.2. Sampling area

2.3. Sample collection

2.4. Grouping of animals

2.5. Fecal Egg Count (FEC)

2.6. Therapeutic trails

2.7. Statistical analysis

3. Result

3.1. Effect of drugs on reduction of EPG on various days in caprine and ovine

4. Discussion

Conclusion

Declarations

References

Status:

Version 1