Eng3

Jfuas No.1 June 2013

17

A preliminary Study on the Toxicity of Aqueous Extracts of

Some Plants from Darfur against Bulinus Truncatus Snails.

Mohammed yousif Adam1’*, Faysal Sawi Ali’2* , Abd-El-Gabar Nasir Gumma,3*

.1 Department of Science, Faculty of Education for Basic Teachers, University of Alfashir,Sudan.

.2 Department of Biology, Faculty of Education, University of Khartoum, Sudan.

.3 Department of Biology, Faculty of Education, University of Khartoum, Sudan.*Corresponding author:[email protected]

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Abstract

In this study, some folk-medicinal used plants from Darfur region

of the Sudan were hypothesized to contain ingredients useful in

control of schistosomiasis intermediate host snails. Twenty seven

plant species were tested for their aqueous extracts molluscicidal

activity against Bulinus truncatus adult snails. The primary screening

الفارش األول-التطبيقيةالعلوممجلة–جامعة م2013يونيو–العدد

18

technique was used. Eight species produced extracts that showed some

degree of molluscicidal activity in bioassays.The molluscicidal

activity of five of them were recorded for the first time in the Sudan.

These were: Alternanthera nodiflora (Amaranthaceae); Amaranthus

viridis (Amaranthaceae); Glinus lotoides (Molluginaceae); Khaya

senegalensis (Meliaceae) and Rorippa indica (Bassiaceae).

KEY WORDS: Schistosomiasis, Aqueous extract, plant molluscicde,

Bulinus truncatus

Introduction

Schistosomiasis (Bilharziasis) is a disease infecting various

mammals including man and domestic livestock. It is caused by blood

flukes of the genus Schistosoma (Brown, 1980). The disease endemic

in seventy four countries (WHO, 1985). In the Sudan schistosomiasis

comes after malaria as one of the most important public health

problems (Bella etal., 1980). In Sudan, El –Kheir and El-Tohami

(1979) screened seventy eight plants used in folk –medicines that were

collected from the Nuba Mountains, the Blue Nile Province and

Khartoum Province for molluscicidal activity. Eighteen plants proved

to possess molluscicidal activity. They were active against Bulinus

truncatus and seven of them were also found to be active against

Biomphalaria pfeifferi. Elamin et al., (2005) evaluated the toxic dose

of Balanites aegyptiaca fruit extract when applied naturally against

both Bulinus truncatus and Biomphalaria pfeifferi. LC90 of both types

of the snail was 9 ppm. Howida (2005) screened seeds, stems, and


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leaves of Jatropha aceroides (Euphorbiaceae) plant for their

molluscicidal activity against Bulinus truncatus. The results of the

aqueous extracts revealed that the seeds, were the most active part,

have an LC100 of 100 ppm, while the same concentration of the leaf

killed 80% of the snails. On the other hand, the stem extract showed

LC50 of 100ppm (24h exposure). Molluscicidal properties of

Pulicaria crispa (Forssk.) Oliv. (Asteraceae) have already been

studied in the Sudan. Ali (1997) investigated the potency of aqueous,

as well as acetone, and ethanol extracts of leaves, stem, and flowers

against B. pfeifferi snails. Salha (1999) investigated toxicity of

aqueous and ethanol extracts of leaves, stem, and flowers on Bulinus

truncatus. Somia (1999) reported the activity of P. crispa leaves

aqueous extracts on Lymnaea cailludi snails and anophline larvae with

LC50 and LC100 on concentrations of 1086.4 ppm .Thus

investigation of molluscicidal activity of indigenous plants in the

control of the snail intermediate host of schistosomiasis in the Sudan

is needed, Darfur has a very rich and varied flora, which has been little

studied. This flora represents an enormous reservoir of potential

molluscicidal activity which is waiting to be discovered. Many of

these plants are used in folk medicine by the people of Darfur to treat

several tropical diseases including schistosomiasis, especially in rural

areas and 1500 ppm of aqueous extracts of leaves respectively. Somia

(2003) also found that juveniles of B. pfeifferi snails were more

susceptible to aqueous extract than adult snails. Atta El Mannan

(2001) showed molluscicidal activity of P. crispa whole plant against


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L. natalensis in the field. Amna (2001) studied the effects of leaves

maturity and storage on the toxicity of P. crispa against Lymnaea

natalensis snails

The objectives of this study is to screen the potentiality of some

plant extracts which may have molluscicidal activities against the

snail Bulinus truncatus intermediate host of schistosmiasis in Darfur.

Materials and Methods

2-1 The Study Area

Alfashir city is the capital of North Darfur State, Western Sudan. It

is situated at latitude ˚13 37 N and longitude ˚25 22 E. It is about 800

Km south west of Khartoum and at an altitude of approximately 740

m above the sea level. The climate is semi-arid with an annual rainfall,

between 100-500 mm. The study was carried out from August 2006 to

September2007.

2-2 Collection and Preparation of Plant Samples

Selection of the plant samples used in this study was based on data

collected from the folk-medicine in Darfur area, assuming that it can

also lead to the discovery of promising new plant molluscicides.

Accordingly, tewenty seven of folk-medicinal plant samples were

collected from Valleys and Goz lands around Alfashir city during the

rainy season (from August-September 2006), when the plants were

flowering and fruiting. The local names, the habitat, methods of

preparation, application and the disease they are claimed to cure were

obtained from local herbalist and recorded (Table 1). The collected


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plants were protected from direct sunlight by covering with wet sacks,

and then transported to the laboratory in University of Alfashir, where,

the collected plants were put separately. Each plant was assorted by

part (leaves, stem, roots, flowers and fruits). Every plant part was air

dried under shade in the laboratory for three to four days. Thereafter,

the plant parts were ground in a wooden mortar with a pestle. The

powdered plant material was then put in labeled air tight plastic bags

and kept in desiccators for further use.

Herbarium specimens of these herbs were kept and transported to

Khartoum for identification. The plant specimens used in this study

were identified by Dr. Abd-El-Gabar Nasir Gumma, a taxonomist

from Biology Department, Faculty of Education, University of

Khartoum and deposited in the laboratory of Faculty of Education for

basic level teachers, University of Alfashir.

2-3 Collection and Maintenance of Snail Samples

Snails were collected from Golo reservoir. The snails were

collected in the early morning (at 8.00-10.00 a.m.) using a flat dip-net

scoop (Demian and Kamel, 1972; Richie etal., 1962). The collected

snails were put in a plastic bucket containing some dam water with

few leaves. The snails were then transported to the laboratory for

identification and maintenance. The snails used in this study were

Bulinus truncatus. In the laboratory, snails were identified to the

species level using the snail identification key published by Danish

Bilharzias Laboratory (DBL, 1983), and were confirmed by National

Research Centre of Khartoum (Plate1). Healthy snails were


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maintained in plastic troughs (12cm depth×30cm diameter) with a

capacity of about 6 liters (Plate2). The trough bottoms were covered

with a layer of clay and some gravel, had been sterilized by heating to

150˚C for at least 18 hours. Tap water used was strongly aerated for

about three days to allow evaporation of chlorine and then, the troughs

filled up to two thirds. Green Eruka sativa(Girgeer) leaves were

immersed in boiling water for about one minute and then cooled in tap

water. After the removal of the mid-rib, leaves were dried and ground.

The powdered dry Girgeer was used for feeding the snails three times

a week. It should be noted that the soft part of the Girgeer leaves were

the most suitable material for feeding the snail. Snails fed on dry ribs

of the leaves did not survive. The aquaria were maintained at

temperature between 25-30˚C. Water was changed twice a month or

when necessary. The dead snails were removed as soon as possible

from the troughs to prevent water fouling.

Plate (1): Bulinus truncatus adult snail Plate (2): B.

truncatus breeding aquarium


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2-4 Preparation of the Aqueous Extracts

Five grams of each powdered dry plant part material (leaves, stem,

roots, flowers, fruits and seeds) of the twenty seven plant species used

in the screening tests were weighted using an electric balance. The

weighted powdered dry parts were soaked in 150 ml of distilled water

for 24 hours with occasional vigorous shaking, using a magnetic

stirrer during the first six hours. Then, the suspension was filtered

using (Whatman No.1) filter paper.The marc was washed with several

portions of distilled water to adjust the volume of the stock solution,

using volumetric flasks to 200 ml (25000ppm).The plant extracts were

used immediately after the extraction to gurantee their freshness.

2-5 Primary Screening Technique

Different stock solutions (5gm/200ml) of the aqueous extracts for

the different parts of the twenty seven plant species were tested

against adult Bulinus truncatus snails according to the method

recommended by WHO (1961). The working solutions were prepared

by using the simple dilution procedure, by adding 1, 3, 5, 7, 9, 11, 13,

15, 17, 19 and 21ml of stock solution to dechlorinated tap water in

containers (10cm depth×25cm diameter) and the total volume adjusted

to 500ml to give concentrations of 50, 150, 250, 350, 450, 550, 650,

750, 850, 950 and 1050 ppm respectively.

Ten healthy, uniform sized adult snails of B. truncatus were put in

each container. The exposure period was 24 hours followed with the

recovery period of 24 hours. For the recovery period, snails were

transferred to other containers with 500ml of dechlorinated tap water


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and fed with powdered dry Girgeer. Dead snails were counted. All the

snails which at the end of the recovery period remained completely

within their shells, floated upside -down on the surface of the solution

or settled at the bottom without response to mechanical prodding were

considered dead. When all the snails were killed, further dilutions

were prepared to determine the least lethal concentration that killed

100% of the snails (LC100), using the simple dilution procedure.

Control tests were reared in similar conditions without treatment. All

treated and control groups were put under artificial light for about

eight hours daily. The surviving snails were removed to other

containers with 500ml of dechlorinated tap water, fed on Girgeer and

observed for three days.

Results

Aqueous Extracts of different parts (leaves, stem, flowers, seeds

and fruits) of twenty seven plant species suggested as medicinal plants

by the local people in Darfur region were screened for their

molluscicidal activity against the snail Bulinus truncatus. They

classified into eighteen families, with Amaranthaceae and

Caesalpiniaceae being the most represented families within the

collection. Out of the tested plant samples eight species were active

against B. truncatus snails as listed in Table (1). Table (2) summarizes

the results of the bioassays, listing only the species that showed some

molluscicidal activity in at least one bioassay.

The lethal doses that caused 100% mortality (LC100) of the adult

B. truncatus snails, when 5gm of the powdered plant species parts


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were soaked in 200 ml of distilled water for 24 hours were:

Alternanthera nodiflora (Amaranthaceae) (leaves 187.36 ppm and

196.22 ppm for the flowers); Glinus lotoides (Molluginaceae) (leaves

736.24 ppm and 684.11 ppm for the fruits); Khaya senegalensis

(Meliaceae) stem bark 531.21 ppm; Rorippa indica (Bassicaceae)

stem 946.16ppm; Amaranthus viridis (Amaranthaceae) leaves 1032.08

ppm; Blanites aegyptiaca (Balanitaceae) fruits 9.42 ppm; Acacia

nilotica (Mimosaceae) fruits 112.50 ppm and Pulicaria crispa

(Asteraceae) leaves 1378.60 ppm.

Table (1): Screened plant species and their local uses

Scientific Name

(Species)Family Local Name

Active

Morphologi

cal Part

Folk-medicine

Uses

1 Conyza

aegyptiaca

Asteraceae Raihan

Elgrouf

Leaves &

stem

Smoke was used as

insects repellant

2 Pulicaria

crispa

Asteraceae Altagar(Rab

oul)

Leaves Hypertension

3 Glinus

lotoides

Molluginaceae Remta

ghabsha

Leaves&

seeds

Drop tape worms

& treating dysuria

4 Alternanther

a nodiflora

Amaranthaceae Gebbis

(Abu-Tagia)

Leaves &

flowers

Treating dysuria &

schistosomiasis

5 Amarathus

spinosa

Amaranthaceae Khadiga

kroa

Leaves Swallows &

treating tonsillitis

6 Amaranthus

viridis

Amaranthaceae Lisan Eltair

Kabeer

Leaves Swallows &

treating tonsillitis

7 Aerua

javanica

Amaranthaceae Ras –Al-

shaeb

Flowers Treating swallows

8 Cassia

obtusifolia

Caesalpiniaceae Kaual Seeds Treating hepatic

infections(specially bile)

9 Senna italica Caesalpiniaceae Sana Kalib Fruits &

roots

Laxative & dermal

diseases treating

10 Cassia

occidentalio

Caesalpiniaceae Sim-

Eldabeib

Stem &

roots

Anti-toxic

11 Senna

alexandrina

Caesalpiniaceae Sanamakka Fruits&

roots

Laxative and

treating hepatic

diseases


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12 Euphorbia

egyptiaca

Euphorbiaceae Um-Libaina

Masrea

Stem& the

milky juice

Treating wounds

13 Euphorbia

hirta

Euphorbiaceae Um-

libaina(Tam

r Elghanam)

Roots Anti toxics

especially of the

scorpion

14 Rorippa

indica

Bassicaceae Hussain

Sakaran

(kamounElgobn)

Stem Analgesic &killing

tape worms

15 Geigaria

alata

Cruciferae Gadgad Stem& roots Treating of

diabetus mellitus&

stomachaches

16 Indigofera

arrecta

Papilionaceae Remta

Hamraa

Stem& roots Dysuria

17 Rogera

adenophylla

Pedaliaceae Teber

Elgadaul

(Um –ganato)

Stem&

seeds

Pertusis

18 Indigofera

hochstetteri

Papilionaceae Dahaseer Stem Toxic

19 Ipomoea

cordofana

Convalvulaceae Ladouk Stem Toxic

20 Zizyphus

spina-christi

Rhamnaceae Sedder Leaves

Fruits

Laxative, drop

worms

21 Aristolochia

bracteolata

Aristolochiaceae Um-Galagil

(ErrgElagrab)

Stem& roots Anti Malaria,

dysentery&hepatic diseases

22 Solanum

dubium

Solanaceae Gibbain Fruits Guardiasis &

making yogurt

23 Trianthema

pentandra

Ficoidaceae Rabaa Stem Anti scorpion

venom

24 Heliotropiu

m

aegyptium

Boranginaceae Danab

Elagrab

roots Treating wounds

25 Khaya

senegalensis

Meliaceae Mahogany Stem bark Treating jaundice,

hook worms

&dermatomes

26 Accacianilotica

Mimosaceae Garadh Fruits Stomach aches,colds&

rheumatism

27 Balanites

aegyptiaca

Balanitaceae Heglieg Fruits Anathematic &

against hepatitis


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Table (2): Plants that Showed Molluscicidal Activity in the Screening

Tests.No. Plant species Family Active Part LC100 (ppm)

1 Acacia nilotica Mimosaceae - Fruits 112.50

2 Alternanthera

nodiflora

Amaranthaceae -Leaves

-Flowers

187.36

196.22

3 Amaranthus viridis Amaranthaceae - Leaves 1032.08

4 Balanites aegyptiaca Balanitaceae -Fruits 9.42

5 Glinus lotoides Molluginaceae -Leaves

-Fruits

736.24

684.11

6 Khaya senegalensis Meliaceae - Stem Bark 531.21

7 Pulicaria crispa Asteraceae - Leaves 1378.60

8 Rorippa indica Bassicaceae - Stem 946.16

Discussion

The deciding factor in schistosomiasis control programmes using

molluscicides is the cost. Synthetic molluscicides are expensive and in

addition, may lead to problems of toxicity to non-target organisms and

deleterious long-term effects on the environment. The possible

development of resistance in schistosomiasis transmitting snails is

another important factor. Other methods of snails control such as

biological methods or drying of the canals also proved unsatisfactory.

Therefore, these considerations urge the use molluscicides of plant

origin that are selectively active, easily biodegradable, inexpensive

and available in endemic areas (WHO, 1983). The success of plant

molluscicidal plans may become in the near future a useful

complement for the control of schistosomiasis.

In the present study, twenty seven folk-medicinal plant species

from Darfur were preliminary screened by part for their molluscicidal

activity against B. truncatus adult snails using the aqueous extract


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method. Eight of them (29.60%) caused a significant death of B.

truncatus adult snails after 24 hours of exposure. This is considered a

good rate, reinforcing the importance of ethno- medical information in

the search of bioactive extracts. Among the eight plant species that

showed molluscicidal activity three have been tested for their

mollscicidal activities by other researchers. Accacia nilotica

(Mimosaceae) was reported by Mardufa and Ouma, (1977); El Kheir

and El Tohami, (1979); Ayoub, (1985). Balanites aegyptiaca

(Balanitaceae) was reported by Archibald, (1933); Wagner, (1936);

Bashir et al., (1984); Elamin et al., (2005). Pulicaria crispa

(Asteraceae) was studied by many researchers (Ali, 1997; Somia,

1999 and 2003; Salha, 1999; Atta El Mannan, 2001; Amna, 2001 and

Ali, 2007). In the present study P. crispa showed less activity against

B. truncatus adult snails in the screening tests than mentioned in

previous studies. These differences may be due to the extraction

methods or to seasonal and geographical variations of the plant.

The molluscicidal activity of Alternanthera nodiflora

(Amaranthaceae); Amaranthus viridis (Amaranthaceae); Glinus

lotoides (Molluginaceae); Khaya senegalensis (Meliaceae) and

Rorippa indica (Bassicaceae) were reported for the first time in the

Sudan (Table1). Further studies are needed to test the potency of

solvent extracts on adult, one day old juveniles, and one day old egg

masses of B. truncatus and other intermediate snail hosts of

schistosomiasis.


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On conclusion, according to the results obtained above, some of

the active species mentioned in this study were considered promising

molluscicides, in control of schistosomiasis. These results need to be

followed by further investigations for comprehensive evaluation and

possible use of the active species in schistosomiasis control

programmes.

Acknowledgement

The authors thanks and appreciation is to Dr. Abdelazziz Abd-

El-Rahim, Director of Schistosomiasis Research Laboratory,

University of Khartoum for his help and advice.

References

Ali, A. E. (1997). Investigation into the molluscicidal activity of

Pulicaria crispa (Forssk.) Oliv. (Compositae) on Biomphalaria

pfeifferi snail vectors of schistosoma mansoni in the Sudan. M. Sc.

Thesis. Faculty of Veterinary Sciences, University of Khartoum,

Sudan.

Amna, H.I. (2001). The Effect of Leaves Maturity and Storage on the

Toxicity of Pulicaria crispa (Forssk.) Oliv. (Compositae) on Lymnaea

natalensis snails. M. Sc. Thesis. Faculty of Education, U. of K.,

Sudan.

Archibald, R. G. (1933). The endemiology and epidemiology of

schistosomiasis in the Sudan. Journal of Tropical Medicine and

Hygiene, 36: 345-348.


30

Ayad, N. (1956). Bilharziasis survey in British Somalia, Eriteria,

Sudan and Yemen, Bulletin of World Health Organization, 14: 1-117.

Atta El-Mannan, A. M. (2001) . Studies on biological control of

Lymnaea natalensis snails using a molluscicidal indigenous plant

Pulicaria crispa (Forssk.) Oliv. (Compositae). Ph. D. Thesis. Faculty

of Veterinary Sciences, University of Khartoum, Sudan

Ayoub, S. M. (1985). Molluscicidal Properties of Acacia nilotica.

Medica, 46: 181-183.

Bashir, A. K. ; Ahmed, G. H. M. ; Suliaman, S. M. and El-Kheir,

Y. M. (1984). Molluscicidal and other biological activities of

Balanites aegyptiaca. The First Arab Conference on Medicinal Plants.

Cairo, Egypt.

Bella, H.; Marschall, T. E. Dec.; A. H. S. and Vaughan, J. P.

(1980). Migrant workers and schistosomiasis in the Gezira,Sudan.

Transactions of Royal Society of Tropical Medicine and Hygiene, 74:

36-39.

Brown, D. S. (1980). Freshwater snails in Africa and their medical

importance. Taylor and Francis Ltd., London.

Danish Biharziasis Laboratory (1983). A field guide to freshwater

snails in countries of the WHO Eastern Mediterranean Region. WHO

Collaborating Centre for Applied Malacology, Copenhagen

Demian, E. S. & Kamel, E. G. (1972). Growth and population

dynamics of Bulinus truncatus under semi –field conditions in Egypt.

Proceeding of Egyptian Academy of Science, 25: 37-60.


31

Elamin, E.; Subki, H. and Koko, W. S. (2005). Utilization of

Balanites aegyptiaca fruit extract in the eradication of schistosomiasis

transmitting snails in Sudan (Primary investigation). Albuhuth Sudan

Journal of Scientific Research 8(2): 112-119.

El- Khier, Y. M. and El- Tohami, M. S. (1979). Investigation of

Molluscicidal Activity of Certain Sudanese plants Used in Folk-

Medicine 1, ll. The London School of Hygiene and Tropical Medicine.

82 (11/12): 237-241.

Howida, A. H. (2005). Investigation on the molluscicidal activity of

Jatropha aceroides against Bulinus truncatus. Albuhuth- Sudan

Journal of Scientific Research, 9 (12): 33-40.

Jurberg, P.; Vasconcellos M. C. and Mendes, N. M. (1989). Plants

empregadas como moluscicidas: Uma visHo critica. Mem. Inst.

Oswaldo Cruz. 84: 76-83.

Kloos, H. and McCullough, F. S. (1987). Plants with recognized

molluscicidal activity. In Mott, K. E. ed. Plant Molluscicides.

Kuo, Y. H. (1987). Plant molluscicidal Studies in the Peoples

Republic of China. In : Mott, K.E. ed. Plant Molluscicides. New York.

UNDP/World Bank/WHO, p.229-298.

Mardufa, A. and Ouma, J. H. (1977). A new Chalcone as a natural

product from Polygonum senegalensis. Phytochemistry, 17: 823-824.

Molgaad, P.; Chihaka, A.; Furu, P.; Ndekha, A. and Nyazema, N.

(1999). Community based schistosomiasis control in Zimbabwe using

the molluscicidal berries of Pytolacca dodencandra AETFAT

conference in Harare, Zimbabwe.


32

Richie, L. S.; Radke, M. E. and Ferguson, F. F. (1962). Population

dynamics of Australorbis glabratus in Puerto Rico. (bid, 11): 171.

Salha, B. F. (1999). The effect of Pulicaria crispa (Forssk). Oliv.

(Compositae) on Bulinus truncatus snail vector of Schistosoma

haematobium in the Sudan. M. Sc. Thesis. Faculty of Education,

University of Khartoum, Sudan.

Somia, A. M. (1999). The effect of Pulicaria crispa (Forssk). Oliv.

(Compositae) on Lymnaea cailludi snails and other aquatic lives. M.

Sc. Thesis. Faculty of Education, University of Khartoum, Sudan.

Somia, A. M. (2003). Ovicidal and Molluscicidal Potency of

Pulicaria crispa (Forssk). Oliv. on Biomphalaria pfeifferi egg-masses,

juveniles and adults. Ph. D. Thesis. Faculty of Education, University

of Khartoum, Sudan.

Wagner, V. A. (1936). The possibility of eradicating bilharzia by

intensive planting of the tree Balanities. South Africa Med. J. 10: 10-

11.

World Health Organization (WHO) (1961). Molluscicides: second

report of expert committee on Bilhaziasis. WHO Technical Report,

Series No. 214

ــــــــــــــــــ��ـ (1983). Report of scientific working group on plant

molluscicide and guidelines for evaluation of plant molluscicide.

Geneva, (TDR/SCH- SWE (4)/ 83.3).

ــــــــــــــــــ��ـ (1985). The control of schistosomiasis. WHO Technical

Report, Series No. 728, Geneva.

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