|Year : 2015 | Volume
| Issue : 2 | Page : 107-114
Effect of nitroxynil (fasciolid) on adult Fasciola gigantica and Fasciola hepatica in infected cows
Eman K Omran, Noha S Ahmad
Department of Medical Parasitology, Faculty of Medicine, Sohag University, Sohag, Egypt
|Date of Submission||12-Dec-2014|
|Date of Acceptance||25-May-2015|
|Date of Web Publication||27-Jan-2016|
Eman K Omran
PhD, Department of Medical Parasitology, Faculty of Medicine, Sohag University, Sohag 2630
Source of Support: None, Conflict of Interest: None
Drug resistance to treatment of fascioliasis with triclabendazole (TCBZ) has emerged in Sohag Governorate, Egypt. Nitroxynil belongs to the halogenated phenol group of fasciolicides. It is highly active against adult liver flukes. A nitroxynil metabolite is produced in the liver parenchyma adding to its flukicidal activity and augmenting its efficacy against late immature flukes that migrate through the liver tissues. Treatment with nitroxynil may be an effective replacement for therapy with TCBZ in cases of resistance.
The aim of this study was to evaluate the efficacy of nitroxynil in the treatment of fascioliasis by assessing its effect on teguments and durability of adult Fasciola gigantica and Fasciola hepatica worms.
Materials and methods
Infected cows were selected on the basis of clinical signs, and infection was confirmed by detection of Fasciola eggs in their stools. Nitroxynil was administered as recommended in two doses 15 days apart, and the animals were slaughtered 15 days after treatment. Fasciola worms collected from the bile ducts were identified and prepared for electron microscopy. Tegument changes were examined with scanning electron microscopy.
The removed adult flukes of both species were moving sluggishly and appeared pale with no evidence of gut content. Scanning electron microscopy examination of these flukes revealed evidence of swelling of the tegument that showed regional variation in its severity. Loss of spines was also observed.
The present study demonstrated the flukicidal properties of nitroxynil, proving that the tegument is an important target for its action. Disruption of the fluke's main line of defense allowed the drug access to other internal tissues, leading to more widespread damage. Nitroxynil may be successfully used for treatment in case of resistance to TCBZ.
Keywords: adult Fasciola, in vivo treatment, flukicides, nitroxynil (flukicide), scanning electron microscope
|How to cite this article:|
Omran EK, Ahmad NS. Effect of nitroxynil (fasciolid) on adult Fasciola gigantica and Fasciola hepatica in infected cows. Parasitol United J 2015;8:107-14
|How to cite this URL:|
Omran EK, Ahmad NS. Effect of nitroxynil (fasciolid) on adult Fasciola gigantica and Fasciola hepatica in infected cows. Parasitol United J [serial online] 2015 [cited 2018 Sep 26];8:107-14. Available from: http://www.new.puj.eg.net/text.asp?2015/8/2/107/175008
| Introduction|| |
Fascioliasis is one of the most important parasitic diseases in domestic ruminants throughout the world caused by the trematodes Fasciola gigantica and Fasciola hepatica. Both flukes are economically significant parasites in livestock constituting one of the important causes of zoonotic infections. In Egypt, fascioliasis has been recorded as an important clinical problem, particularly among school-aged children living in rural areas of the Nile Delta , . Animal as well as human fascioliasis is a growing problem, as it has been recorded in almost all governorates, especially those of the Nile Delta in Lower Egypt  . A large variety of animals, such as sheep, goats, cattle, buffalo, horses, donkeys, camels, and rabbits, showed Fasciola infection rates that may reach 90% in some areas  . Among these animal species, fascioliasis is highly endemic in sheep as indicated by macroscopic detection of liver flukes in slaughtered sheep during abattoir surveys (20.6%)  . In another report from North Sinai, 12.7% of Fasciola spp. were recorded in sheep and goats by microscopic detection of eggs  . In the same year, infection was found to occur mostly in sheep (17.84%), followed by cows (12.31%), buffaloes (9.73%), and lastly goats (5.40%)  . Reported prevalence rates for Fasciola infections have reduced in recent years due to control measures enforced in the Egyptian governorates, including triclabendazole (TCBZ) administration  .
In the absence of any effective vaccines, the principal method of control remains to be the use of drugs. Treatment with TCBZ, which is a member of the benzimidazole family of anthelminthics, is the main therapy for control of fascioliasis morbidity  . It works by preventing the polymerization of the tubulin molecule into the cytoskeletal microtubule structures , and is particularly active against both juvenile and adult flukes  . Likewise, closantel, nitroxynil, rafoxanide, and TCBZ are among the few flukicides that are active against both mature and immature liver flukes. Other flukicides such as albendazole, clorsulon, and oxyclozanide are active against adult liver fluke only  .
However, resistance of F. hepatica to TCBZ has been registered since 1990  and has since been reported in Australia  , Ireland  , the Netherlands  , and southern Wales  . It is now known to exist in a number of European countries as well , , and in Egypt  . A number of fasciolicides (including nitroxynil) have been evaluated against TCBZ-resistant flukes  .
Nitroxynil (4-hydroxy-3-iodo-5-nitrobenzonitrile), which is a halogenated phenol , , acts by stopping oxidative phosphorylation in the cell mitochondria. This disturbs the production of ATP, thus impairing motility of the parasites , . Nitroxynil binds very strongly (97-98%) to plasma proteins  , indicating that oral ingestion is the most likely route. Its nematocidal activity against adult and larval stages of Haemonchus contortus in sheep and Haemonchus placei, Oesophagostomum radiatum, and Bunostomum phlebotomum in cattle was previously recorded  . It was reportedly active against TCBZ-resistant F. hepatica  and was found synergistically active when therapy was tested in combination with clorsulon or closantel in the treatment of salicylanilide-resistant flukes, even when administered in lower than recommended doses  . Therapy with nitroxynil gave 100% efficacy against Fasciola spp. in all post-treatment observations , . Efficacies around 100% have also been described under field  and experimental  conditions.
The integrity of the surface plasma membrane and the tegumental syncytium is essential as a first line of defense for flukes. The tegument is the primary drug target immediately exposed to anthelminthics. In 2010, Toner et al.  observed that, due to the severity of changes produced by TCBZ treatment, the flukes become eliminated from the host through the gall bladder and that their migration corresponds with the degenerative changes. In their detailed report, they described progressive disruption of the tegumental system and musculature, which became severe with time of exposure, up to complete sloughing of the tegument and degeneration of the underlying tissues. Disruption of the gastrodermis also increased in severity over time, although it was not as severely affected as the tegument. They added that, notably, the recovered flukes displayed disruption of the subtegumental muscle bundles that appeared widely separated, as well as progressive loss of fibers that rendered the muscle blocks almost empty and barely recognizable  . It was noted that, on lengthening the period of treatment with 40 mg/kg nitroxynil, the activity of the adult fluke was affected up to immobility  . The earliest tegumental changes were recorded after 24 h from onset of treatment.
Most recently, in 2015, Junquera  stated that, in contrast to other anthelminthics (e.g. imidazothiazoles, benzimidazoles, and tetrahydropyrimidines), nitroxynil has a residual effect i.e., it not only kills the parasites present in the host at the time of treatment, but also protects against reinfestation for a period of up to several weeks, depending on the dose and the specific parasite. This further collaborates the former declaration that nitroxynil can be used as an alternative to TCBZ in therapy of resistant cases  .
Scanning electron microscopy (SEM) has been successfully used for evaluating anthelminthic effect on changes of tegumental surface of flukes. SEM allows scanning of the whole-body surface before examination with transmission electron microscope of internal structure changes  , revealing the location and possible variation of any disruption caused by a drug  . Therefore, the aim of the present study was to evaluate the efficacy of nitroxynil for the treatment F. gigantica and F. hepatica in naturally infected cows, by noting its action on the tegument and durability after treatment.
| Materials and methods|| |
Type of study
This is a clinical veterinary trial.
Animal source of flukes
Infected cows were selected from Tahta Veterinary Clinic on the basis of clinical signs such as foul-smelling watery diarrhea, anorexia, and hypothermia with abrupt decrease in milk yield. Natural infection was determined by detection of Fasciola eggs in the cows' stools using direct smear and sedimentation methods.
Nitroxynil 1 ml/25 kg body weight was given orally to two naturally infected cows according to the instructions of Chemical Industries Development under license of Pharmachim Bulgaria. Two doses were given 15 days apart in an attempt to avoid any side effects.
Flukes identified as adult F. gigantica and F. hepatica were recovered from the bile ducts of slaughtered cows 15 days after treatment and washed in several changes of warm (37°C) sterile physiological saline to remove any attached particles.
Scanning electron microscopy studies
Preparation of samples for SEM was carried out as described  . Washed flukes were fixed in 5% glutaraldehyde for 24-72 h and washed four times in sodium cacodylate buffer (pH 7.3) for about 15 min. Each postfixation was carried out by adding 1% osmium tetroxide for 2 h and then the samples were washed three times in the same buffer. Dehydration was carried out in ascending concentrations of ethanol (30%, 50%, 70%, and 90%), each for 30 min, and in double changes of absolute ethanol for 24-48 h. The samples were incubated at 20-25°C on double-sided carbon scotch tape and coated with gold. The samples were prepared, examined, and photographed at the SEM Unit, Assuit University.
| Results|| |
Flukes recovered from the bile ducts of cows 15 days after treatment were moving sluggishly and appeared pale with no evidence of gut contents. The SEM examination revealed evidence of swelling of the tegumental surfaces, with regional variation in the severity of the swelling. On the ventral surfaces of the flukes, moderate swelling of the teguments was observed around the oral and ventral suckers ([Figure 1] and [Figure 2]). In the central regions of the oral cones, the teguments were severely swollen. The swelling was less severe along the lateral margins of the oral cones. At the same time, the spines were barely visible ([Figure 3] and [Figure 4]). In the anterior mid-body regions, directly behind the ventral suckers, no spines were visible as they had become completely submerged by the externally swollen teguments ([Figure 5] and [Figure 6]). The mid-body regions displayed widespread swelling and extensive furrowing of the teguments. The spines were almost submerged by the surrounding swollen teguments ([Figure 7] and [Figure 8]). The dorsal surfaces were similarly acutely affected as the ventral surfaces. The spines appeared to be flaking off, and there were multiple pores and furrowing of the teguments ([Figure 9] and [Figure 10]).
|Figure 1: SEM of F. gigantica after treatment in vivo with Nitroxynil 1 cc/25 kg BW. Ventral Surface: The apical cone and the anterior mid-body regions revealed tegumental swelling with regional variations in severity surrounding the oral and ventral suckers (arrow).|
Click here to view
|Figure 2: SEM of F. gigantica after treatment in vivo with Nitroxynil 1 cc/25 kg BW. Ventral Surface: In the central region of the oral cone the tegument was severely swollen and the spines were barely visible (arrows).|
Click here to view
|Figure 3: SEM of F. gigantica after treatment in vivo with Nitroxynil 1 cc/25 kg BW. Ventral Surface: In the anterior mid-body region, directly behind the ventral sucker, no spines were visible as they had become completely submerged by the extremely swollen tegument.|
Click here to view
|Figure 4: SEM of F. gigantica after treatment in vivo with Nitroxynil 1 cc/25 kg BW. Ventral Surface: In the mid-body region, the spines appeared almost submerged by the swollen tegument surrounding them.|
Click here to view
|Figure 5: SEM of F. gigantica after treatment in vivo with Nitroxynil 1 cc/25 kg BW. Dorsal Surface: The spines appeared to be fl aking off, with multiple pores and furrowing of the tegument (arrows).|
Click here to view
|Figure 6: Identified SEM of F. hepatica after treatment in vivo with Nitroxynil 1 cc/25 kg BW. Ventral surface. Tegumental swelling of the apical cone, showed regional variations in severity surrounding the oral and ventral suckers (arrows).|
Click here to view
|Figure 7: SEM of F. hepatica after treatment in vivo with Nitroxynil 1 cc/25 kg BW. Ventral surface. Identified area between the oral and ventral sucker showed displaced spines (arrow).|
Click here to view
|Figure 8: SEM of F. hepatica after treatment in vivo with Nitroxynil 1 cc/25 kg BW. Ventral surface. The mid-body region laterally showed displaced spines directly behind the ventral sucker, no spines were visible as they had become completely submerged by the extremely swollen tegument.|
Click here to view
|Figure 9: SEM of F. hepatica after treatment in vivo with Nitroxynil 1 cc/25 kg BW. Ventral surface. The central mid-body region showed severe swelling and furrowing of the tegument (arrow).|
Click here to view
|Figure 10: SEM of F. hepatica after treatment in vivo with Nitroxynil 1 cc/25 kg BW. Dorsal surface. The spines appeared to be fl aking off with multiple pores and furrowing of the tegument (arrow).|
Click here to view
| Discussion|| |
Treatment of animal fascioliasis is imperative for the management of economic losses and for avoiding zoonotic infections. Cows and sheep pose as the main hosts for both Fasciola spp. and are responsible for passing the infection to man  . The effective control of these infections with anthelminthic drugs is therefore vital for both animal welfare and productivity ,, . The drug of choice in the treatment of fascioliasis has been TCBZ, until the flukes started to exhibit their resistance to it  . Resistance of liver flukes to treatment with nitroxynil has not reached the scale experienced with nematodes, but it has been reported for salicylanilides, rafoxanide, and closantel, with cross resistance to nitroxynil. Another report on treatment with closantel and nitroxynil was published in SCOPS  .
It was postulated that the administration of these flukicides to animals used for human consumption can lead to transfer of drug residues to human food ,, . In 2013, all drug products containing the active ingredients of clorsulon, closantel, nitroxynil, rafoxanide, and TCBZ were prohibited for use in lactating animals producing milk for human consumption. The reason presented was that residues in undetectable concentrations in milk could be concentrated during manufacturing, and become detectable in milk products  . The researchers recommended treatment of dairy cows during the dry period, when no milk is being produced for human consumption, and advised that an adequate withdrawal period is allowed for the elimination of drug residues before resumption of milking.
In vitro studies have shown great drug disruption of worms following treatment with clorsulon and nitroxynil , . Besides ingestion through the gut, the tegument presents an absorptive surface for the uptake of drugs by the fluke. The adult fluke is bathed in bile when in the bile ducts of the host, which are its preferential dwelling site, and becomes exposed to drugs excreted in the flow of bile. Thus, because the tegument is immediately exposed to anthelminthics, it is a crucial drug target. The drug-induced damage of the tegument facilitates drug penetration to deeper-lying tissues, thus leading to serious effects on the flukes. It was deduced by McKinstry et al.  that, in the in vivo situation, the surfactant action of bile and the immune response aggravates the damage, leading to the discoloration of the flukes observed after 72 h treatment. Nitroxynil is excreted mostly unchanged through the liver giving rise to high concentrations in the bile ducts. Recent research showed that a metabolite of nitroxynil produced in the liver parenchyma added to its flukicidal activity and explained its efficacy against late immature flukes that migrate through the liver tissues  .
In the present study, the flukes recovered from the bile ducts of nitroxynil-treated cows 15 days after treatment were not dead and moved sluggishly. They appeared pale with no evidence of gut contents. As described, healthy flukes are grayish-brown in color, with a flattened spiny tegument, through which the dark outline of blood-filled ceca is evident  . The pallor and empty guts of flukes, observed in our study confirms that penetration and autophagia of substances such as bile and drugs takes place  . As regards other flukicides, it was reported that praziquantel produces flaccid paralysis of the fluke, starting with initial increase followed by decrease in muscle tone  . The researchers added that oxyclozanide, rafoxanide, niclofolan, bithionol, and hexachlorophene also induce rapid spastic paralysis of the fluke. Nitroxynil produces a similar effect, which explains the sluggish movement recorded in our study. By discontinuing oxidative phosphorylation in the cell mitochondria, it disrupts the production of ATP, thus causing a long-term suppression of movement , . In another report, the flukes after treatment in vivo with nitroxynil showed little or no sign of movement and appeared yellow in color, with no evidence of gut contents. Swelling of the mid-body was visible to the naked eye  .
The present study has confirmed that nitroxynil causes swelling and severe disruption of the tegumental surface of Fasciola, as well as swelling of the basal in-folds in the tegumental syncytium, edematous flooding of the parenchyma, and mitochondrial deformation in the syncytium and tegumental cells , . In our study, no spines were visible in the anterior mid-body region, directly behind the ventral sucker, as they had become completely submerged by the externally swollen tegument. The mid-body region also displayed widespread swelling and extensive furrowing of the tegument, and the spines were almost submerged by the swollen tegument surrounding them. From previous studies, disruption to the spines seems to be a particular feature of nitroxynil action ,, . When due to the action of the drug, the thin tegument covering the spines splits and sloughs off and the spines are lost, the holes thus formed in the syncytium allow more access of the drug to internal tissues. This becomes especially promoted by the surfactant action of bile in vivo  . The cause for disruption of spine structure observed in our study and reported with other fasciolicides is unknown ,, .
In the present study, the dorsal surface disruption was as severe as that seen on the ventral surface. The spines appeared to be flaking off, and multiple pores and furrowing of the tegument were detected. This outcome of in vivo treatment with nitroxynil was previously defined as regional differences in tegumental disruption that is more severe on the dorsal than on the ventral surface, and on the anterior than on the posterior region of the fluke  . In their investigation, the researchers also described extensive swelling and blebbing of the tegument on both surfaces, and at high magnification microvillus-like projections were evident with some tegumental loss in the oral region of the fluke. In our point of view, greater affection of the dorsal surface resulted from higher exposure to the drug as the flukes are folded ventrally when in the bile ducts.
McKinstry et al.  stated that, after 24 h of in vivo treatment with 40 mg/kg of nitroxynil, the recovered flukes were moving normally but appeared pale, with no evidence of gut contents. After 48 h, the flukes were less active, and surface swelling could be observed. After 72 h, the flukes showed little or no sign of movement; all appeared yellow with empty gut contents. Swelling of the mid-body was visible to the naked eye. In comparison, another report found that after the full course of in vivo treatment with TCBZ the recovered flukes remained active for 48 h displaying limited morphological disruption, but they were all dead after 72 h. The researchers described some blebbing and sloughing of the tegument around the oral sucker, when an extra layer had apparently been deposited on the fluke surface, giving it a flattened appearance  . A relatively recent investigation indicated that each of TCBZ-susceptible and TCBZ-resistant isolates were affected by nitroxynil treatment in vitro and in vivo, showing ultrastructural changes similar to those previously seen in the Cullompton TCBZ-susceptible isolate  . In the present study, 15 days after treatment the flukes were still alive, but suffered severe swelling of the tegument, accompanied by isolated areas of flattening along the lateral margins of the flukes and in the tail region. Our SEM examination of nitroxynil-treated flukes showed limited areas of tegument sloughing in the tail region. In more seriously affected specimens, the syncytium appeared stripped away to reveal the basal lamina. Some deeper lesions were also observed as the loss of tegument facilitated penetration of the drug to the exposed deeper tissues of the flukes. There was disruption of the subtegumental tissues with spacing or flooding between the cells and breakdown of the cell bodies, to the extent that often they were difficult to identify. Similar findings were described  in TCBZ-treated flukes. The flooding feature was also recorded in other drug studies on F. hepatica using sulfoxide metabolite of the compound alpha  and metabolites of TCBZ as sulfoxide (TCBZSO) and sulfone (TCBZSO 2 )  . A very recent publication in 2015  revealed that, in sheep with a high fluke burden, TCBZ was ineffective in treating chronic fasciolosis indicating TCBZ resistance. At the same time nitroxynil and closantel proved to be wholly active against TCBZ-resistant flukes in chronically infected sheep with high chronic fluke burdens.
In conclusion, our study has confirmed previous SEM studies with nitroxynil in that the drug effectively disrupts the tegument of Fasciola. In addition, it causes disruption to the gut following in vivo treatment, an observation that is consistent with the oral uptake of the drug as well. Giving similar therapeutic results with little resistance, nitroxynil therapy of fascioliasis in rural animals may be considered a safe replacement for TCBZ treatment.
Author contribution: EK Omran and NS Ahmad proposed the research idea, conducted the work, and shared in writing and revision of the manuscript.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Esteban JG, Gonzalez C, Curtale F, Muñoz-Antoli C, Valero MA, Bargues MD, et al
. Hyperendemic fascioliasis associated with schistosomiasis in villages in the Nile Delta of Egypt. Am J Trop Med Hyg 2003; 69:429-437.
El-Shazly AM, El-Beshbishi SN, Azab MS, El-Malky M, Abdeltawab AH, Morsy AT. Past and present situation of human fascioliasis in Dakahlia Governorate, Egypt. J Egypt Soc Parasitol 2009; 39:247-262.
Soliman MF. Epidemiological review of human and animal fascioliasis in Egypt. J Infect Dev Ctries 2008; 2:182-189.
Farag HF. Human fascioliasis in some countries of the Eastern Mediterranean Region. East Mediterr Health J 1998; 4:156-160.
El-Shazly AM, Abdel-Magied AA, El-Nahas HA, El-Metwaly MS, Morsy TA, El Sharkawy EM, Morsy AT. On the main reservoir host of Fasciola
in Dakahlia Governorate, Egypt. J Egypt Soc Parasitol 2005; 35:243-252.
Mazyad SA, el-Nemr HI. The endoparasites of sheep and goats, and shepherd in North Sinai Governorate, Egypt. J Egypt Soc Parasitol 2002; 32:119-126.
El-Shazly AM, Haridy SF, Soliman M, Rifaat MMA, Morsy TA. Fascioliasis among live and slaughtered animals in nine centers of Dakahlia Governorate. J Egypt Soc Parasitol 2002; 32:47-57.
Keiser J, Engels D, Büscher G, Utzinger J. Triclabendazole for the treatment of fascioliasis and paragonimiasis. Expert Opin Investig Drugs 2005; 14:1513-1526.
Fairweather I. Triclabendazole: new skills to unravel an old(ish) enigma. J Helminthol 2005; 79:227-234.
Fairweather I. Triclabendazole progress report, 2005-2009: an advancement of learning? J Helminthol; 2009, 83:139-150.
Boray JC, Crowfoot PD, Strong MB, Allison JR, Schellenbaum M, Von Orelli M, et al
. Treatment of immature and mature Fasciola hepatica
infections in sheep with triclabendazole. Vet Rec 1983; 113:315-317.
Power C, Sayers R, O'Brien B, Furey A, Danaher M, Jordan K. Review of studies on flukicide residues in cows' milk and their transfer to dairy products.
Irish J Agric Food Res 2013; 52:197-207.
Brennan GP, Fairweather I, Trudgett A, Hoey E, McCoy, McConville M, et al
. Understanding triclabendazole resistance. Exp Mol Pathol 2007; 82:104-109.
Overend DJ, Bowen FL. Resistance of Fasciola hepatica
to triclabendazole. Aust Vet J 1995; 72:275-276.
Mulcahy G, Dalton JP. Vaccines in control of liver fluke infections in ruminants: current status and prospects. Irish Vet J 1998; 51:520-525.
Moll L, Gaasenbeek CP, Vellema P, Borgsteede FH. Resistance of F. hepatica
against triclabendazole in cattle and sheep in the Netherlands. Vet Parasitol 2000; 24:153-158.
Thomas L, Goles GC, Duuffus K. Triclabendazole resistance F. hepatica
in South-Western Wales. Vet Rec 2002; 12:200-209.
Alvarez-Sánchez MA, Mainar-Jaime RC, Pérez-García J, Rojo-Vázquez FA. Resistance of Fasciola hepatica
to triclabendazole and albendazole in sheep in Spain. Vet Rec 2006; 159:424-425.
Shalaby HA, El Namaky AH, Kamel ROA. In vitro
effect of artemether and triclabendazole on adult Fasciola gigantica
. Vet Parasitol 2009; 160:76-82.
Borgsteede FHM, Moll L, Vellema P, Gaasenbeek CPH. Lack of reversion in triclabendazole resistant F. hepatica
. Vet Rec 2005; 156:350-351.
Whelan M, Kinsella B, Furey A, Moloney M, Cantwell H, Lehotay SJ, Danaher M. Determination of anthelmintic drug residues in milk using ultra high performance liquid chromatography-tandem mass spectrometry with rapid polarity switching. J Chromatogr A 2010; 1217:4612-4622.
Ghoneim MM, El-Ries M, Hassanein AM, Abd-Elaziz AM. Voltammetric assay of the anthelminthic veterinary drug nitroxynil in bulk form and formulation at a mercury electrode. J Pharm Biomed Anal 2006; 41:1268-1273.
Boray JC, Happich A. Standardized hemotherapeutical tests for immature and mature F. hepatica
infections in sheep. Aust Vet J 1968; 44:72-78.
Rapic D, Dzakula N, Sakar D, Richards RJ. Comparative efficacy of triclabendazole, nitroxynil and rafoxanide against immature and mature F. hepatica
in naturally infected Cattle. Vet Rec 1998; 122:59-62.
Alvinerie M, Floc'h R, Galtier P. Plasma protein binding of nitroxynil in several species. J Vet Pharmacol Ther 1991; 14:170-173.
Martin RJ. Modes of action of anthelmintic drugs. Vet J 1997; 154:11-34.
Coles GC, Stafford KA. The activity of oxyclozanide, nitroxynil, clorsulon and albendazole against adult triclabendazole resistant F. hepatica
. Vet Rec 2001; 148:723-724.
Fairweather I, Boray JC. Fasciolicides: efficacy, actions, resistance and its management. Vet J 1999; 158:81-112.
Wellington AC. Nitroxynil. Anthelmintic activity in cattle following subcutaneous injection. J S Afr Vet Assoc 1978; 49:125-126.
Keyyu JD, Kassuku AA, Kyvsgaard NC, Monrad J. Comparative efficacy of anthelmintics against Fasciola gigantica
and amphistomes in naturally infected cattle in Kilolo district, Tanzania. Tanzan Vet J 2008; 25:40-47.
31. Mooney L, Good B, Hanrahan JP, Mulcahy G, de Waal T. The comparative efficacy of four anthelmintics against a natural acquired Fasciola hepatica
infection in hill sheep flock in the west of Ireland. Vet Parasitol 2009; 164:201-205.
Toner E, Brennan GP, Hanna RE, Edgar HW, Fairweather I. Time-dependent changes to the tegumental system and gastrodermis of adult Fasciola hepatica
following treatment in vivo
with triclabendazole in the sheep host. Vet Parasitol 2010; 174:218-227.
McKinstry B, Halferty L, Brennan GP, Forbes AB. F. hepatica
tegumental surface alterations following treatment in vivo
and In vitro
with nitroxynil (Trodax). Parasitol Res 2003; 91:251-263.
Junquera P. Nitroxinil
-nitroxynil for veterinary use on cattle, sheep and goats against flukes and roundworms. Parasitipedia. net 2014;19:37.
Martínez-Valladares M, Famularo M, Fernández-Pato N, Castañón-Ordóñez L, Cordero-Pérez C, Rojo-Vázquez FA. Efficacy of nitroxynil against F. hepatica
resistant to triclabendazole in a naturally infected sheep flock. Parasit Res 2010; 107:1205-1211.
McKinstry B, Brennan GP, Halferty L, Forbes AB, Fairweather I. Ultrastructural changes induced in the tegument and gut of F. hepatica
after in vivo
and in vitro
drug treatment with nitroxynil (Trodax). Parasitol Res 2007; 101:929-941.
Hayat MA. Principles and technique of electron microscopy
, 2nd ed. New Jersey: University Park Press; 1981. 1.
Boray JC. Fascioliasis. In: Handbook Series in Zoonoses
. Section C. Parasitic Zoonoses. Hillyer GV & Hopla CE edit, CRC Press, Boca Raton, Florida 1982;3:71-88.
McKellar QA, Jackson F. Veterinary anthelmintics: old and new. Trends Parasitol 2004; 20:456-461.
Rehman MA, Samad MA. Pattern of occurrence of single and concurrent diseases associated with mortality in commercial chickens in Bangladesh. Bangl J Vet Med 2003; 1:15-20.
SCOPS. Sustainable control of parasites in sheep. 2012. Available at: http://www.scops.org.Uk\endoparasite-Liver-fluke.html
Imperiale F, Ortiz P, Cabrera M, Farias C, Sallovitz JM, Iezzi S, et al.
Residual concentrations of the flukicidal compound triclabendazole in dairy cows' milk and cheese. Food Addit Contam Part A Chem Anal Control Expo Risk Assess 2011; 28:438-445.
Meaney M, Fairweather I, Brennan GP, McDowell LSL, Forbes AB. F. hepatica
: Effects of the fasciolicide clorsulon in vitro
and in vivo
on the tegumental surface, and a comparison of the effects on young- and old-mature flukes. Parasitol Res 2003; 91:238-250.
Ichhpujani RL, Bhatia R. Medical parasitology
. 3rd ed. New Delhi, India: Jaypee Brothers Medical Publishers; 2002. 242.
Fairweather I, Holmes SD, Threadgold LT. Fasciola hepatica
: motility response to fasciolicides In vitro
. Exp Parasitol 1984; 57:209-224.
McKinstry B. Ultrastructural changes observed in Fasciola hepatica
following treatment with nitroxynil and triclabendazole, alone and in combination [PhD thesis]. UK: the Queen's University of Belfast; 2008.
McKinstry B, Halferty L, Brennan GP, Fairweather I. Morphological response of triclabendazole-susceptible and triclabendazole resistant isolates of F. hepatica
to treatment in vitro
with nitroxynil (Trodax). Parasitol Res 2009; 104:645-655.
Meaney M, Fairweather I, Brennan GP, Forbes AB. Transmission electron microscope study of the ultrastructural changes induced in the tegument and gut of Fasciola hepatica
following in vivo
drug treatment with clorsulon. Parasitol Res 2004; 92:232-241.
Meaney M, Allister J, McKinstry B, McLaughlin K, Brennan GP, Forbes AB, Fairweather I. Fasciola hepatica
: ultrastructural effects of a combination of triclabendazole and clorsulon against mature fluke. Parasitol Res 2007; 100:1091-1104.
McConville M, Brennan GP, McCoy M, Castillo R, Hernandez-Campos A, Ibarra F, Fairweather I. Adult triclabendazole-resistant Fasciola hepatica
: surface and subsurface tegumental responses to in vitro
treatment with the sulphoxide metabolite of the experimental fasciolicide compound alpha. Parasitology 2006; 133(Pt 2):195-208.
Halferty L, Brennan GP, Hanna RE, Edgar HW, Meaney MM, McConville M, et al
. Tegumental surface changes in juvenile Fasciola hepatica
in response to treatment in vivo
with triclabendazole. Vet Parasitol 2008; 155:49-58.
Halferty L, Brennan GP, Trudgett A, Hoey L, Fairweather I, Relative activity of triclabendazole metabolites against the liver fluke F. hepatica.
Vet Parasitol 2009; 159:126-138.
Hanna RE, McMahon C, Ellison S, Edgar HW, Kajugu PE, Gordon A, et al
. Fasciola hepatica
: a comparative survey of adult fluke resistance to triclabendazole, nitroxynil and closantel on selected upland and lowland sheep farms in Northern Ireland using faecal egg counting, coproantigen ELISA testing and fluke histology. Vet Parasitol 2015; 207:34-43.
[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7], [Figure 8], [Figure 9], [Figure 10]