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The predatory capability of Arthrobotrys cladodes var. macroides in the control of Haemonchus contortus infective larvae
Veterinary Parasitology 130 (2005) 263–266 www.elsevier.com/locate/vetpar
The predatory capability of Arthrobotrys cladodes var. macroides in the control of Haemonchus contortus infective larvae A. Eslami a,*, S. Ranjbar-Bahadori b, R. Zare c, M. Razzaghi-Abyaneh d a
Department of Parasitology, Faculty of Veterinary Medicine, Tehran University, P.O. Box 14155-6453 Tehran, Iran b Faculty of Veterinary Medicine, Azad Islamic University of Garmsar, Garmsar, Iran c Department of Botany, Plant Pests & Diseases Research Institute, Tehran, Iran d Department of Mycology, Pasteur Institute of Iran, Tehran 13164, Iran Received 23 October 2004; received in revised form 30 March 2005; accepted 30 March 2005
Abstract One hundred compost samples were examined for the presence of nematophagous fungi on the sheep farms of Mazanderan, province, Iran. Arthrobotrys cladodes var. macroides (IRAN 677C = CBS 143565) was isolated from 3% of the samples examined. Nematophagous activity of this fungus which was shown for the first time in this study, revealed the addition of 1000, 8000, 20 000 and 100 000 conidia per gram of feces of sheep reduced significantly (P < 0.001) the number of Haemonchus contortus infective larvae in the feces by 41.71%, 63, 27%, 73.49% and 94.96%, respectively. These results show that A. cladodes var. macroides is a promising candidate for biological control of H. contortus. # 2005 Elsevier B.V. All rights reserved. Keywords: Arthrobotrys cladodes var. macroides; Biological control; Haemonchus contortus
1. Introduction Gastrointestinal nematodes of sheep and goats constitute a major problem in sheep industry in Iran (Eslami and Nabavi, 1976) and their control increase considerably production (Skerman et al., 1967). In addition to traditional chemical anthelmintic drugs, new alternative control strategies, specially the use of * Corresponding author. Tel.: +98 21 6924469; fax: +98 21 6933222. E-mail address: [email protected] (A. Eslami).
trapping fungi as natural enemies have been investigated for controlling ruminant parasitic nematodes during the past decades (Gronvold et al., 1987; Larsen et al., 1991). Among 70 genera and 160 fungal species associated with nematodes (Duddington, 1994; Waller and Faedo, 1996), more than 50 species are able to capture and kill nematodes in soil (Siddiqui and Mahmood, 1996). This study was designed to isolate nematophagous fungi from compost samples and to show their suitability in biological control of L3 of Haemonchus contortus.
0304-4017/$ – see front matter # 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.vetpar.2005.03.039
264
A. Eslami et al. / Veterinary Parasitology 130 (2005) 263–266
2. Materials and methods 2.1. Soil samples Over a period of one year (2001–2002), 100 compost samples were obtained from different sheep farms of Mazandaran province (north of Iran along the Caspian sea coast), with moist and moderate climatic conditions. These were examined for the presence of nematophagous fungi. 2.2. Fungi To isolate the nematophagous fungi, 5 g of each compost sample was mixed with 50 ml sterile water in a 250 ml Erlenmeyer flask by shaking thoroughly and was kept at room temperature (25–30 8C) for 30 min. Then, 5 ml of the supernatant was mixed with 0.5 ml antibiotic (200 unit penicillin and 400 mg streptomycin) and left at room temperature for 24 h. It was then centrifuged for 15 min at 1500 rpm. Primary isolation of nematophagous fungi was achieved by using chloramphenicol–2% water agar (CHF–WA) medium. This medium consisted of chloramphenicol (0.05% dissolved in ethanol), Difco agar (2%) and demineralized water (1 L). All ingredients were mixed in Erlenmeyer flask and divided into 10 cm diameter Petri dishes. After autoclaving for 15 min at 121 8C, CHF–WA was also used for subsequent isolation and subculturing of the fungi. The fungal isolates were preserved on potato dextrose agar (PDA, Difco).
fecal culture. To obtain third stage larvae (L3) of H. contortus, the fecal cultures were placed on a Baermann funnel for 12 h to collect the larvae. 2.5. Experimental procedure Surface of the cultures was washed thoroughly with 2 ml of demineralized water for each plate using a sterile glass rode and the number of conidia was calculated using a haemocytometer slide under the microscope. Four groups each of three sheep fecal cultures were separately seeded each with 1000, 8000, 20 000 and 100 000 conidia of Arthrobotrys cladodes var. macroides in 1 ml demineralized water. Demineralized water free of fungal conidia was added to control plates. Fecal cultures were maintained on plastic trays and incubated at 25 8C for 7 days. Then each plate was placed on a Baermann funnel for 12 h in order to collect the remaining larvae. Total number of larvae was calculated from 10 aliquots (0.05 ml each) placed on microscopic slides and observed using a streomicroscope. 2.6. Statistics Analysis of variance using one-way ANOVA was used to compare the means of the groups. Also, post hoc test: Tukey was applied to show the relationship between increasing numbers of fungal conidia and reduction in parasite larvae.
2.3. Haemonchus contortus egg donor
3. Results
A 12-month old female native sheep free of gastrointestinal parasitic nematodes was orally infected with 5000 H. contortus infective larvae. During the experiment, the sheep was fed manually. Feces collected directly from the rectum were homogenized and the mean number of H. contortus egg per gram of feces (EPG) was estimated (1052).
The fungus was identified A. cladodes var. macroides according to Van Oorschot (1985). Morphologically, fungal conidiophores were erect, mostly simple, some producing one to two side branches, up to 320 mm long, not proliferating, conidiogenous heads irregularly swollen, bearing conidia on narrow denticles. Conidia elongate-obovoidal to elongateellipsoidal, measuring 15–25 mm 6–11 mm, 1-septate near middle, immature conidia 1-celled. Chlamidospores were present. This fungus was the only nematophagous fungus identified and isolated from 3% of the compost samples collected from Mazandaran province of Iran. Its nematophagous activity which was shown for the first time in the present study
2.4. Fecal culture Fifty grams of homogenized sheep feces containing H. contortus eggs were cultivated in special glass jars with 10 cm diameter at 25 8C for 7 days. Demineralized water was added to each jar to keep wet the
A. Eslami et al. / Veterinary Parasitology 130 (2005) 263–266
Fig. 1. Predatory activity of Arthrobotrys cladodes var. macroides against Haemonchus contortus by producing special traps around the infective larvae (L3). Table 1 Nematophagous activity of Arthrobotrys cladodes var. macroides in vitro Number of conidia (per gram of feces)
Recovered larvae (L3) (mean S.D.)
L3 reduction (%)
1000 8000 20000 100000 Control (without conidia)
400 27.2 252 35.1 182 31.8 35 2.38 686 47.5
41.71 63.27 73.44 94.96 0
was highly performed by forming traps around the H. contortus third stage larvae in their environment (Fig. 1). The number of larvae from the fecal culture and the percentage reduction with respect to the original inoculum are shown in Table 1. The reduction values of H. contortus infective larvae in each batch of conidia in comparison with control group was highly significant (P < 0.001) and also between increasing added numbers of conidia and reduction in parasite larvae (P < 0.05) except those of 8000 and 20 000 conidia. In general, the highest and the lowest predatory activity (94.96% and 41.71%) were achieved by adding 100 000 and 1000 conidia per gram of feces, respectively.
4. Discussion Most of the investigation carried out on the predatory activities of Arthrobotrys are dealing with
265
the specimens donated by other research laboratories or investigators (Waller and Faedo, 1993; Pandey, 1973; Gronvold et al., 1987). A few reports exist on nematophagous activity of native isolates (Sanyal, 2000; Chandrawathani et al., 2001; Manueli et al., 1999). This survey achieved the objective of identifying a native and rare isolate of Arthrobotrys, e.g. A. cladodes var. macroides from compost samples. Although it was previously reported as nematophagous fungi (1944) and some isolates: CBS 499.99 collected by Ysk (1990) from soil Banana plantation from Tenerif Spain and CBS 120.54 collected by Peach (1954) are deposited at culture collection of this Bureau, yet no other reports have been found on its predatory activity on ruminant parasites. A. cladodes var. macroides showed a high individual predatory activity against H. contortus infective larvae in the present study. To compare our results with those of other species of Arthrobotrys, our findings are in line with many other workers (Mendoza-De and Vazquez-Prats, 1994; Gronvold et al., 1985; Waller and Faedo, 1993). Our findings showed nematophagous activity of A cladodes var. macroides has a direct relationship with the number of conidia added. The highest and the lowest reduction in the number of L3 of H. contortus were achieved by adding 100 000 (94.96%) and 1000 (41.71%) conidia, respectively. These results are partly similar with the findings of Mendoza-De and Vazquez-Prats (1994) with Monacrosporium eudermatum. Addition of 20 000 and 100 000 conidia per gram of feces reduced 95.7% and 100% of infective larvae of H. contortus. Bird and Herd (1995) also showed a reduction of 40.5% and 95.8% in the number of infective cyathostome larvae from naturally infected horses by adding 1 and 100 spores of A. oligospora, respectively. This is not always a rule. Because addition of 20 000 (Mendoza-De and Vazquez-Prats, 1994), 2500 (Gronvold et al., 1985) and 20 (Hashmi and Connan, 1989) conidia per gram of feces infected with H. contortus and Cooperia sp. and Ostertagia ostertagi, and C. oncophora induced 98.3%, 99% and 99% and 72% reduction in the number of larvae, respectively. Similarities and differences in nematophagous activities of A. cladodes var. macroides with that of other workers seems to be natural. As it was stated by Mendoza-De and Vazquez-Prats (1994) even there is a marked differences in predatory activity between
266
A. Eslami et al. / Veterinary Parasitology 130 (2005) 263–266
A. oligospora strains, because some strains need only a few conidia to get the expected effect whereas other strains need a large inoculum to get the same effect. Results in the present paper have shown that fungus A. cladodes var. macroides has a satisfactory predatory activity when directly added to sheep feces; thus, it is a potential biological control agent of gastrointestinal parasitic nematodes in sheep. Further investigations are needed to show the in vivo passage of this fungus through gastrointestinal tract of sheep.
Acknowledgements The authors wish to thank Research Council of Tehran University for financial support of this project (Grant no. 215/3/515) and Dr. W. Gams (CBS, the Netherlands) for confirming the identity of the fungus used in this study.
References Bird, J., Herd, R.P., 1995. In vitro assessment of two species of nematophagous fungi (Arthrobotrys oligospora and Arthrobotrys flagrans) to control the development of infective cyathostome larvae from naturally infected horses. Vet. Parasitol. 36, 181–186. Chandrawathani, P., Hoglund, J., Waller, P.J., Jamnah, O., 2001. Prospects for controlling small ruminant nematodes by predacious fungi: survey, isolation and identification of a Malaysian isolate for biological control of helminths. In: Second International Congress/13th VAM Congress and CVA-Australasia Regional Symposium, 27–30 August 2001, Kuala Lumpur, pp. 125–126. Duddington, C.L., 1994. Predacious fungi and nematode 1962. Experientia 50, 414–420. Eslami, A., Nabavi, L., 1976. Species of gastrointestinal nematodes of sheep from Iran. Bull. Soc. Pathol. Exot. 96, 92–95. Gronvold, J., Korsholm, H., Wolstrup, J., Nansen, P., Henriksen, S.A., 1985. Laboratory experiments to evaluate the ability of
Arthrobotrys oligospora to destroy infective larvae of Cooperia species and to investigate the effect of physical factors on the growth of the fungus. J. Helminthol. 59, 119–125. Gronvold, J., Wolstrup, J., Henriksen, S.A., Nansen, P., 1987. Field experiment on the ability of Arthrobotrys oligospora (Hyphomycetales) to reduce the number of larvae of Cooperia oncophora (Trichostrongylidae) in cow pats and surrounding grass. J. Helminthol. 61, 65–71. Hashmi, H.A., Connan, R.M., 1989. Biological control of ruminant trichostronglids by Arthrobotrys oligospora, a predacious fungus. Parasitol. Today 5, 28–30. Larsen, M., Wolstrup, J., Henriksen, S.A., Dackman, C., Gronvold, J., Nansen, P., 1991. In vitro stress selection of nematophagous fungi for biocontrol of parasitic nematodes in ruminants. J. Helminthol. 65, 193–200. Manueli, P.R., Waller, P.J., Faedo, M., Mahommed, F., 1999. Biological control of nematode parasites of livestock in Fiji: screening of fresh dung of small ruminants for the presence of nematophagous fungi. Vet. Parasitol. 81, 39–45. Mendoza-De, G.P., Vazquez-Prats, V.M., 1994. Reduction of Haemonchus contortus infective larvae by three nematophagous fungi in sheep fecal culture. Vet. Parasitol. 55, 197– 203. Pandey, V.S., 1973. Predatory activity of nematode trapping fungi against the larvae of Thichostrongylus axei and Ostertagia ostertagi: a possible method for biological control. J. Helminthol. 11, 35–48. Sanyal, P.K., 2000. Screening for Indian isolates of predacious fungi for use in biological control against nematode parasite of ruminants. Vet. Res. Commun. 24, 55–62. Siddiqui, Z.A., Mahmood, I., 1996. Biological control of plant parasitic nematodes by fungi: a review. Bioresour. Technol. 58, 229–239. Skerman, K.O., Shahlapoor, A.A., Eslami, A., Eliazian, M., 1967. Observation on the incidence, epidemiology, control and economic importance of gastrointestinal parasites of sheep and goats in Iran. Vet. Med. Rev. 141–152. Van Oorschot, C.A.N., 1985. Taxonomy of Dactylaria complex V. A review of Arthrobotrys and allied genera. Stud. Mycol. 26, 61– 96. Waller, P.J., Faedo, M., 1993. The potential of nematophagous fungi to control the free-living stages of nematode parasites of sheep: Screening studies. Vet. Parasitol. 49, 285–297. Waller, P.J., Faedo, M., 1996. The prospects for biological control of the free-living stages of nematode parasites of livestock. Int. J. Parasitol. 26, 915–925.
The predatory capability of Arthrobotrys cladodes var. macroides in the control of Haemonchus contortus infective larvae A. Eslami a,*, S. Ranjbar-Bahadori b, R. Zare c, M. Razzaghi-Abyaneh d a
Department of Parasitology, Faculty of Veterinary Medicine, Tehran University, P.O. Box 14155-6453 Tehran, Iran b Faculty of Veterinary Medicine, Azad Islamic University of Garmsar, Garmsar, Iran c Department of Botany, Plant Pests & Diseases Research Institute, Tehran, Iran d Department of Mycology, Pasteur Institute of Iran, Tehran 13164, Iran Received 23 October 2004; received in revised form 30 March 2005; accepted 30 March 2005
Abstract One hundred compost samples were examined for the presence of nematophagous fungi on the sheep farms of Mazanderan, province, Iran. Arthrobotrys cladodes var. macroides (IRAN 677C = CBS 143565) was isolated from 3% of the samples examined. Nematophagous activity of this fungus which was shown for the first time in this study, revealed the addition of 1000, 8000, 20 000 and 100 000 conidia per gram of feces of sheep reduced significantly (P < 0.001) the number of Haemonchus contortus infective larvae in the feces by 41.71%, 63, 27%, 73.49% and 94.96%, respectively. These results show that A. cladodes var. macroides is a promising candidate for biological control of H. contortus. # 2005 Elsevier B.V. All rights reserved. Keywords: Arthrobotrys cladodes var. macroides; Biological control; Haemonchus contortus
1. Introduction Gastrointestinal nematodes of sheep and goats constitute a major problem in sheep industry in Iran (Eslami and Nabavi, 1976) and their control increase considerably production (Skerman et al., 1967). In addition to traditional chemical anthelmintic drugs, new alternative control strategies, specially the use of * Corresponding author. Tel.: +98 21 6924469; fax: +98 21 6933222. E-mail address: [email protected] (A. Eslami).
trapping fungi as natural enemies have been investigated for controlling ruminant parasitic nematodes during the past decades (Gronvold et al., 1987; Larsen et al., 1991). Among 70 genera and 160 fungal species associated with nematodes (Duddington, 1994; Waller and Faedo, 1996), more than 50 species are able to capture and kill nematodes in soil (Siddiqui and Mahmood, 1996). This study was designed to isolate nematophagous fungi from compost samples and to show their suitability in biological control of L3 of Haemonchus contortus.
0304-4017/$ – see front matter # 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.vetpar.2005.03.039
264
A. Eslami et al. / Veterinary Parasitology 130 (2005) 263–266
2. Materials and methods 2.1. Soil samples Over a period of one year (2001–2002), 100 compost samples were obtained from different sheep farms of Mazandaran province (north of Iran along the Caspian sea coast), with moist and moderate climatic conditions. These were examined for the presence of nematophagous fungi. 2.2. Fungi To isolate the nematophagous fungi, 5 g of each compost sample was mixed with 50 ml sterile water in a 250 ml Erlenmeyer flask by shaking thoroughly and was kept at room temperature (25–30 8C) for 30 min. Then, 5 ml of the supernatant was mixed with 0.5 ml antibiotic (200 unit penicillin and 400 mg streptomycin) and left at room temperature for 24 h. It was then centrifuged for 15 min at 1500 rpm. Primary isolation of nematophagous fungi was achieved by using chloramphenicol–2% water agar (CHF–WA) medium. This medium consisted of chloramphenicol (0.05% dissolved in ethanol), Difco agar (2%) and demineralized water (1 L). All ingredients were mixed in Erlenmeyer flask and divided into 10 cm diameter Petri dishes. After autoclaving for 15 min at 121 8C, CHF–WA was also used for subsequent isolation and subculturing of the fungi. The fungal isolates were preserved on potato dextrose agar (PDA, Difco).
fecal culture. To obtain third stage larvae (L3) of H. contortus, the fecal cultures were placed on a Baermann funnel for 12 h to collect the larvae. 2.5. Experimental procedure Surface of the cultures was washed thoroughly with 2 ml of demineralized water for each plate using a sterile glass rode and the number of conidia was calculated using a haemocytometer slide under the microscope. Four groups each of three sheep fecal cultures were separately seeded each with 1000, 8000, 20 000 and 100 000 conidia of Arthrobotrys cladodes var. macroides in 1 ml demineralized water. Demineralized water free of fungal conidia was added to control plates. Fecal cultures were maintained on plastic trays and incubated at 25 8C for 7 days. Then each plate was placed on a Baermann funnel for 12 h in order to collect the remaining larvae. Total number of larvae was calculated from 10 aliquots (0.05 ml each) placed on microscopic slides and observed using a streomicroscope. 2.6. Statistics Analysis of variance using one-way ANOVA was used to compare the means of the groups. Also, post hoc test: Tukey was applied to show the relationship between increasing numbers of fungal conidia and reduction in parasite larvae.
2.3. Haemonchus contortus egg donor
3. Results
A 12-month old female native sheep free of gastrointestinal parasitic nematodes was orally infected with 5000 H. contortus infective larvae. During the experiment, the sheep was fed manually. Feces collected directly from the rectum were homogenized and the mean number of H. contortus egg per gram of feces (EPG) was estimated (1052).
The fungus was identified A. cladodes var. macroides according to Van Oorschot (1985). Morphologically, fungal conidiophores were erect, mostly simple, some producing one to two side branches, up to 320 mm long, not proliferating, conidiogenous heads irregularly swollen, bearing conidia on narrow denticles. Conidia elongate-obovoidal to elongateellipsoidal, measuring 15–25 mm 6–11 mm, 1-septate near middle, immature conidia 1-celled. Chlamidospores were present. This fungus was the only nematophagous fungus identified and isolated from 3% of the compost samples collected from Mazandaran province of Iran. Its nematophagous activity which was shown for the first time in the present study
2.4. Fecal culture Fifty grams of homogenized sheep feces containing H. contortus eggs were cultivated in special glass jars with 10 cm diameter at 25 8C for 7 days. Demineralized water was added to each jar to keep wet the
A. Eslami et al. / Veterinary Parasitology 130 (2005) 263–266
Fig. 1. Predatory activity of Arthrobotrys cladodes var. macroides against Haemonchus contortus by producing special traps around the infective larvae (L3). Table 1 Nematophagous activity of Arthrobotrys cladodes var. macroides in vitro Number of conidia (per gram of feces)
Recovered larvae (L3) (mean S.D.)
L3 reduction (%)
1000 8000 20000 100000 Control (without conidia)
400 27.2 252 35.1 182 31.8 35 2.38 686 47.5
41.71 63.27 73.44 94.96 0
was highly performed by forming traps around the H. contortus third stage larvae in their environment (Fig. 1). The number of larvae from the fecal culture and the percentage reduction with respect to the original inoculum are shown in Table 1. The reduction values of H. contortus infective larvae in each batch of conidia in comparison with control group was highly significant (P < 0.001) and also between increasing added numbers of conidia and reduction in parasite larvae (P < 0.05) except those of 8000 and 20 000 conidia. In general, the highest and the lowest predatory activity (94.96% and 41.71%) were achieved by adding 100 000 and 1000 conidia per gram of feces, respectively.
4. Discussion Most of the investigation carried out on the predatory activities of Arthrobotrys are dealing with
265
the specimens donated by other research laboratories or investigators (Waller and Faedo, 1993; Pandey, 1973; Gronvold et al., 1987). A few reports exist on nematophagous activity of native isolates (Sanyal, 2000; Chandrawathani et al., 2001; Manueli et al., 1999). This survey achieved the objective of identifying a native and rare isolate of Arthrobotrys, e.g. A. cladodes var. macroides from compost samples. Although it was previously reported as nematophagous fungi (1944) and some isolates: CBS 499.99 collected by Ysk (1990) from soil Banana plantation from Tenerif Spain and CBS 120.54 collected by Peach (1954) are deposited at culture collection of this Bureau, yet no other reports have been found on its predatory activity on ruminant parasites. A. cladodes var. macroides showed a high individual predatory activity against H. contortus infective larvae in the present study. To compare our results with those of other species of Arthrobotrys, our findings are in line with many other workers (Mendoza-De and Vazquez-Prats, 1994; Gronvold et al., 1985; Waller and Faedo, 1993). Our findings showed nematophagous activity of A cladodes var. macroides has a direct relationship with the number of conidia added. The highest and the lowest reduction in the number of L3 of H. contortus were achieved by adding 100 000 (94.96%) and 1000 (41.71%) conidia, respectively. These results are partly similar with the findings of Mendoza-De and Vazquez-Prats (1994) with Monacrosporium eudermatum. Addition of 20 000 and 100 000 conidia per gram of feces reduced 95.7% and 100% of infective larvae of H. contortus. Bird and Herd (1995) also showed a reduction of 40.5% and 95.8% in the number of infective cyathostome larvae from naturally infected horses by adding 1 and 100 spores of A. oligospora, respectively. This is not always a rule. Because addition of 20 000 (Mendoza-De and Vazquez-Prats, 1994), 2500 (Gronvold et al., 1985) and 20 (Hashmi and Connan, 1989) conidia per gram of feces infected with H. contortus and Cooperia sp. and Ostertagia ostertagi, and C. oncophora induced 98.3%, 99% and 99% and 72% reduction in the number of larvae, respectively. Similarities and differences in nematophagous activities of A. cladodes var. macroides with that of other workers seems to be natural. As it was stated by Mendoza-De and Vazquez-Prats (1994) even there is a marked differences in predatory activity between
266
A. Eslami et al. / Veterinary Parasitology 130 (2005) 263–266
A. oligospora strains, because some strains need only a few conidia to get the expected effect whereas other strains need a large inoculum to get the same effect. Results in the present paper have shown that fungus A. cladodes var. macroides has a satisfactory predatory activity when directly added to sheep feces; thus, it is a potential biological control agent of gastrointestinal parasitic nematodes in sheep. Further investigations are needed to show the in vivo passage of this fungus through gastrointestinal tract of sheep.
Acknowledgements The authors wish to thank Research Council of Tehran University for financial support of this project (Grant no. 215/3/515) and Dr. W. Gams (CBS, the Netherlands) for confirming the identity of the fungus used in this study.
References Bird, J., Herd, R.P., 1995. In vitro assessment of two species of nematophagous fungi (Arthrobotrys oligospora and Arthrobotrys flagrans) to control the development of infective cyathostome larvae from naturally infected horses. Vet. Parasitol. 36, 181–186. Chandrawathani, P., Hoglund, J., Waller, P.J., Jamnah, O., 2001. Prospects for controlling small ruminant nematodes by predacious fungi: survey, isolation and identification of a Malaysian isolate for biological control of helminths. In: Second International Congress/13th VAM Congress and CVA-Australasia Regional Symposium, 27–30 August 2001, Kuala Lumpur, pp. 125–126. Duddington, C.L., 1994. Predacious fungi and nematode 1962. Experientia 50, 414–420. Eslami, A., Nabavi, L., 1976. Species of gastrointestinal nematodes of sheep from Iran. Bull. Soc. Pathol. Exot. 96, 92–95. Gronvold, J., Korsholm, H., Wolstrup, J., Nansen, P., Henriksen, S.A., 1985. Laboratory experiments to evaluate the ability of
Arthrobotrys oligospora to destroy infective larvae of Cooperia species and to investigate the effect of physical factors on the growth of the fungus. J. Helminthol. 59, 119–125. Gronvold, J., Wolstrup, J., Henriksen, S.A., Nansen, P., 1987. Field experiment on the ability of Arthrobotrys oligospora (Hyphomycetales) to reduce the number of larvae of Cooperia oncophora (Trichostrongylidae) in cow pats and surrounding grass. J. Helminthol. 61, 65–71. Hashmi, H.A., Connan, R.M., 1989. Biological control of ruminant trichostronglids by Arthrobotrys oligospora, a predacious fungus. Parasitol. Today 5, 28–30. Larsen, M., Wolstrup, J., Henriksen, S.A., Dackman, C., Gronvold, J., Nansen, P., 1991. In vitro stress selection of nematophagous fungi for biocontrol of parasitic nematodes in ruminants. J. Helminthol. 65, 193–200. Manueli, P.R., Waller, P.J., Faedo, M., Mahommed, F., 1999. Biological control of nematode parasites of livestock in Fiji: screening of fresh dung of small ruminants for the presence of nematophagous fungi. Vet. Parasitol. 81, 39–45. Mendoza-De, G.P., Vazquez-Prats, V.M., 1994. Reduction of Haemonchus contortus infective larvae by three nematophagous fungi in sheep fecal culture. Vet. Parasitol. 55, 197– 203. Pandey, V.S., 1973. Predatory activity of nematode trapping fungi against the larvae of Thichostrongylus axei and Ostertagia ostertagi: a possible method for biological control. J. Helminthol. 11, 35–48. Sanyal, P.K., 2000. Screening for Indian isolates of predacious fungi for use in biological control against nematode parasite of ruminants. Vet. Res. Commun. 24, 55–62. Siddiqui, Z.A., Mahmood, I., 1996. Biological control of plant parasitic nematodes by fungi: a review. Bioresour. Technol. 58, 229–239. Skerman, K.O., Shahlapoor, A.A., Eslami, A., Eliazian, M., 1967. Observation on the incidence, epidemiology, control and economic importance of gastrointestinal parasites of sheep and goats in Iran. Vet. Med. Rev. 141–152. Van Oorschot, C.A.N., 1985. Taxonomy of Dactylaria complex V. A review of Arthrobotrys and allied genera. Stud. Mycol. 26, 61– 96. Waller, P.J., Faedo, M., 1993. The potential of nematophagous fungi to control the free-living stages of nematode parasites of sheep: Screening studies. Vet. Parasitol. 49, 285–297. Waller, P.J., Faedo, M., 1996. The prospects for biological control of the free-living stages of nematode parasites of livestock. Int. J. Parasitol. 26, 915–925.