Sustainable controlofgastrointestinalnematodes (GIN) in small ruminants has been based on the use of alternative methods, including targeted selective treatment, such as FAMACHA. Another GIN control alternative is the use of herbal medicines, although in many cases their use is based on empirical knowledge. Biopolymer nanoformulations has been investigated to maximize the essential oil effects against sheepgastrointestinalnematodes. The aim of the pre- sent study was to combine a Eucalyptus staigeriana essential oil nanoemulsion (EsNano) with FAMACHA as an alternative control for sheep haemonchosis. The study was performed over six months at a commercial sheep farm located in a semiarid region of Northeast Brazil. Initially, a fecal egg count reduction test (FECRT) in sheep with levamisole, ivermectin and oxfendazole in sheep was performed used to determine the most effective anthelmintic to use as the positive control. Levamisole has been selected because it showed efficacy superior to 95%. EsNano was obtained and then its physicochemical properties were characterized. The average (±SE) size of the particles in the nanoemulsion was 276.8 (±12.3) nm with bimodal distribution and polydispersity. Nine visits were performed, from April to September 2013, with an interval of 17 days. One hundred sixty-two male and female sheep were divided into three groups (n=54 each) and were treated when FAMACHA score was 3, 4, or 5: G-EsNano 250mg kg -1 EsNano;
Plant latex, which is normally rich in proteases, has traditionally been used to treat parasitic infections in humans (HANSSON et al., 1986) and animals (CALDWELL & CALDWELL, 1929; SATRIJA et al., 1994), but one of the first reports of the protein nature of the anthelmintic activity was made by Robbins (1930), who identified a cysteine protease named ficin, from Ficus spp., capable of degrading the cuticle of Ascaris suum. A similar effect was observed when A. suum and H. contortus were treated in vitro with bromelain from Ananas comosus (BERGER & ASENJO, 1939; DOMINGUES et al., 2013) and pure crystalline papain from Carica papaya (BERGER & ASENJO, 1940). These proteins quickly led to in vitro ulcerations in the cuticle of the nematode, followed by digestion. However, in vivo evaluation of bromelain on sheep did not reduce H. contortus (DOMINGUES et al., 2013)
The anthelmintic activity of some tanniferous plants are highlighted because condensed tannins are promising compounds in the controlofgastrointestinalnematodes (Suleiman et al. 2014, Yoshihara et al. 2014, Raza et al. 2016). These compounds act directly on the nematodes via formation of complexes with proteins of the helminths, causing damage to the digestive tract, reproductive system and structures such as the cuticle and potentially leading to parasite death (Hoste et al. 2012). These compounds may indirectly contribute to the protein availability for absorption in the small intestine of the animal and provide the best immune response of the host and optimize the antioxidant response to the oxidative stress generated by the parasitic infection (Hoste et al. 2012, Jin et al. 2012, Lopes et al. 2016). Phenolic compounds of natural origin, such as flavonoids and tannins, also play
WHITLOCK 1939). The sheep were divided into four experimental groups containing six animals each. Group I received a single 3 g/10 kg live weight dose of M. thaumasium pellets that had been stored for 36 months. Group II received newly produced M. thaumasium pellets using the same dose and single administration. Group III received newly produced M. thaumasium pellets without fungi using the same dose and single administration. Group IV (control group) did not receive pellets. Coprocultures (ROBERTS and O'SULLIVAN, 1950) of other sheepof the IFPB herd, with an EPG > 1000, were carried out to obtain larvae.
program viz. once in 3 months. Fifty-five naturally infected Madras Red lambs of 6-12 months of age were randomly distributed into four treatment groups of eleven animals each, and the fifth one was main- tained as a control group. Four anthelmintics viz. albendazole (Azole, Vet India, 5.0 mg/kg b.wt), fen- bendazole (Fzole Vet India, 7.0 mg/kg b.wt), levam- isole hydrochloride (Alved, 7.5 mg/kg b.wt) and Zycloz (Closantel, Zydus Agrovet, 10.0 mg/kg b.wt) were used in the study. All the drugs were adminis- tered orally.
The average eosinophil counts were higher in the group treated with homeopathy. On two occasions, there were statistically significant differences between group means. Eosinophilia is a well documented feature of helminth infections but the precise nature of the interaction between parasite and eosinophil remains an enigma (WILDBLOOD et al., 2005). Rainbird et al. (1998) showed that eosinophils can adhere to, and immobilize the invasive L3 larval stage of H. contortus in in vitro cultures in the presence of specific antiparasite antibodies. However, according to Wildblood et al. (2005), ovine nematodes may actively encourage recruitment of eosinophils, since local eosinophil-mediated mucosal damage possibly may provide a permissive local microenvironment for the parasite. Throughout the experiment, it was possible to identify nine naturally-resistant animals (four in the treated and five in the control group) which were not treated individually with conventional anthelmintics. These results agree with several studies that demonstrated that parasite infection resistance is not uniform in the herds. Animals that are resistant, susceptible, or that show intermediary resistance to helminth infections are usually found (BARGER, 1985; AMARANTE et al., 1998). According to Bishop and Stear (2003), modeling studies predict that selective breeding ofsheep for resistance may be used as an effective complementary control strategy for nematode infections with considerable effect in the epidemiology, since it causes reductions in egg output, pasture larval contamination, and, hence, subsequent larval challenge. Therefore, selecting resistant animals seems to be an alternative for a much more promising control for helminth infections than prophylaxis based on treating animals with allopathic or homeopathic drugs.
The anthelmintic resistance stimulated the search for strategies for controlling gastrointestinalnematodes, including the use of free essential oils or its nanoemulsion. This study evaluated the anthelmintic efficacy of Cymbopogon citratus essential oil (CcEO) and C. citratus essential oil nanoemulsion (CcEOn). Pysicochemical analyses were performed. The in vitro effect was determined using the egg hatch test (EHT) on Haemonchus contortus and in vivo effect was evaluated in sheep infected with gastrointestinalnematodes. The animals were treated with CcEO (500 mg/kg) or CcEOn (450 mg/kg) for the fecal egg count (FEC) and the determination of worm burden. The main component of CcEO was citral. The CcEO content in the nanoemulsion was 20% (v/v), and the mean particle size was 248 nm. In EHT, CcEO and CcEOn (1.25 mg/mL) inhibited larval hatching by 98.4 and 97.1%, respectively. Three animals treated with CcEO died whereas in the group treated with CcEOn one animal died. The FEC and total worm burden of the treated groups did not differ from the negative control (p>0.05). The CcEOn showed efficacy only on H. contortus (p<0.05). In conclusion, nanoencapsulation reduced toxicity and increased efficacy on H. contortus.
Gastrointestinal Nematode Infection (GIN) are the main constraint to the production of small ruminants. Studies of medicinal plants have been an important alternative in the effort to control these parasites. Therefore, the purpose of this study was to evaluate the in vitro ovicidal and larvicidal activity of essential oil of Rosmarinus officinalis. The oil was extracted, analyzed by gas chromatography and tested on GIN eggs and larvae in six concentrations, 227.5mg/mL, 113.7mg/mL, 56.8mg/mL, 28.4mg/mL, 14.2mg/mL and 7.1mg/mL. To determine the ovicidal activity, GIN eggs were recovered from sheep feces and incubated for 48h with different concentrations of the oil. For the evaluation of larval migration, third-stage larvae (L3) were obtained by fecal culture, and associated with the essential oil for 24h at the same concentrations, after which they were left for another 24 hours on microsieves, followed by the count of migrating and non-migrating larvae. The assays of R. officinalis oil showed a significant (p<0.05) 97.4% to 100% inhibition of egg hatching and a significant (p<0.05) 20% to 74% inhibition of larval migration. The main constituent revealed by gas chromatography was Eucalyptol. The results indicate that R. officinalis essential oil has ovicidal and larvicidal activity on sheep GINs.
(3/4) farms studied. It is also possible to see that there was a natural increase of EPG of the control groups at all farms, showing that there was no environmental factor associated to the decrease of EPG values in the treated groups during the experimental period. The genera of parasites identified before the treatment of the animals (D-1) are present in Table 2. Nine days after the use of this compound (D+9), there was no significative change in the proportion of the genera ofnematodes identified in the control group of all farms (data not show). However, some changes appear in the treated groups, pointing to different levels of resistance of the nematode population present at each farm (Table 3). It was possible to ascertain on D+9 that some animals of the treated groups at all farms present suggestive clinical signs of persistent infection by gastrointestinalnematodes. Those signs include apathy, lethargy, and anemia of mucous membranes, pointing to a treatment inefficacy.
Ojeda-Robertos NF, Torres-Acosta JF, Aguilar-Caballero AJ, Ayala- Burgos A, Cob-Galera LA, Sandoval-Castro CA, et al. Assessing the efficacy of Duddingtonia flagrans chlamydospores per gram of faeces to control Haemonchus contortus larvae. Vet Parasitol 2008; 158(4): 329- 335. PMid:19028012. http://dx.doi.org/10.1016/j.vetpar.2008.08.022. Ojeda-Robertos NF, Torres-Acosta JF, Mendoza-de-Gives P, Gonzalez- Garduño R, Valero-Coss RO, Liébano-Hernández E, et al. Optimizing the use of Duddingtonia flagrans chlamydospores against Haemonchus contortus in feces ofsheep. Trop Subtrop Agroecosyst 2015; 18: 259-265. Park AW, Haven J, Kaplan R, Gandon S. Refugia and the evolutionary epidemiology of drug resistance. Biol Lett 2015; 11(11): 20150783. PMid:26601680. http://dx.doi.org/10.1098/rsbl.2015.0783. Roeber F, Jex AR, Gasser RB. Impact ofgastrointestinal parasitic nematodesofsheep, and the role of advanced molecular tools for exploring epidemiology and drug resistance - an Australian perspective. Parasit Vectors 2013; 6(1): 153. PMid:23711194. http://dx.doi.org/10.1186/1756-3305-6-153. Sahoo A, Khan F. Nutritional and biological control synergism against gastrointestinalnematodes in small ruminants. J Vet Sci Anim Husb 2016; 4(1): 104.
Medicinal plants are used to control parasites, and their administration is based in ethnoveterinary knowledge. Studies are needed to validate compounds derived from plants that have antiparasitic activity because they are safer and cheaper than commercial anthelmintics, besides to reduce veterinary drug residues in products of animal origin (CHAGAS, 2008; HOSTE et al., 2015). The anthelmintic action of essential oils is associated with major components, justifying the investigation of the anthelmintic action of these compounds for the development of new anthelmintic drugs (MACEDO et al., 2013; RIBEIRO et al., 2013). Thymol is a monoterpene phenol with many pharmaceutical properties including the following: anthelmintic (CAMURÇA-VASCONCELOS et al., 2007), acaricide (ARAÚJO et al., 2015), anti-Leishmania infantum (MORAIS et al. 2014), anti-Trypanosoma cruzi (ESCOBAR et al., 2010) and anti-Echinococcus multilocularis (MARÍA & CELINA, 2014). However, thymol is more toxic than many esters, therefore, the synthesis of TA was performed to obtain a derivative of thymol with improved biological activity and less toxicity (MORAIS et al., 2014). In vitro tests are used to select compounds that have anthelmintic activity (HOUNZANGBE-ADOTE et al., 2005). Thymol showed superior efficacy than TA, probably because the acetylated compound could not penetrate the three layers (outer vitelline, average chitin and inner layer of lipids) that form the H. contortus eggshell (MANSFIELD et al., 1992). The fibrils of chitin in the intermediate layer hinders the penetration of acetylated compounds because the higher liposolubility conferred by the acetyl radical only allows greater penetration in lipid membranes. Carvacryl acetate (CA), a derivate acetylated from carvacrol,
This study aimed to evaluate the anthelmintic activity of Eucalyptus citriodora essential oil and citronellal on sheepgastrointestinalnematodes. Essential oil composition was determined by gas chromatography mass spectrometry. The substances were evaluated in vitro using adult worm motility test (AWMT) and transmission electron microscopy (TEM). The acute toxicity test in mice and the fecal egg count reduction test (FECRT) in sheep were performed. Citronellal was confirmed as the essential oil major constituent (63.9%). According to the AWMT, 2 mg/mL of essential oil and citronellal completely inhibited Haemonchus contortus motility at 6 h post exposure. H. contortus exposed to essential oil and citronellal exhibited internal ultrastructural modifications. The lethal dose 50 values in mice were 5,000 and 2,609 mg/kg for essential oil and citronellal, respectively. E. citriodora essential oil reduced sheep epg at 14 days post treatment by 69.5% (P<0.05). No significant differences were observed in epg between the citronellal and negative control groups (P>0.05). The interaction between citronellal and other constituents in the essential oil may be relevant for its in vivo anthelmintic activity. Thus, E. citriodora essential oil and citronellal pharmacokinetic studies may help elucidate the anthelmintic activity of these compounds.
The large number of studies on AHR in Brazil reflects the concern of the country, which has production potential and seeks to organize the sheep and goat production chain. The vast worldwide literature on AHR in small ruminants contributes to guidelines for controlling the situation. However, this requires an analysis of these studies with factors specific to each case, in order to establish a correlation between the diagnosis and the size of herds, management practices, animal species, parasite species, drugs that are used, and climate. The frequency of use and the maintenance of the effectiveness of anthelmintics over the years are important factors in understanding resistance mechanisms, but are only scantily reported in current studies. It is essential to calculate effectiveness by genus or species of nematode for studies involving combinations of drugs or new antiparasitics. Despite its many limitations, the FECRT is still the most widely used and most feasible test in the country, but should be standardized considering the differences between farms. Considering the high vulnerability of anthelmintics and the scanty prospects for the release of new ones, management practices should not only be evaluated in the diagnosis of AHR but should also be part of parasite control systems. In vitro and molecular effectiveness tests are very promising for the rapid and accurate diagnosis of AHR, but they should be performed with knowledge about the reality in the field to render them feasible for the producer. Molecular studies could be underpinned by analyses of data from field studies, for example, about the maintenance over time of the effectiveness of some antiparasitics such as levamisole in in vivo tests, which suggests that specific studies may contribute to prolong the effectiveness of anthelmintics.
There was a statistically significant difference (p < 0.05) regarding the numbers of treatments with Levamisole Hydrochloride 5% when EPG ≥ 1500. In the fungus + chemical group, only eight deworming treatments were administered, mainly in the early part of the experiment. However, in the chemical group, 17 deworming treatments were necessary, and these were evenly distributed until the end of the experiment. Also in the fungus + chemical group, none of the animals required rescue deworming at any time during the experiment. In the chemical group, two sheep received rescue deworming. In the control group, seven rescue deworming treatments were needed. According to Araújo et al. (1998), clinical parasitism does not occur when nematophagous fungi are administered, because of the diminished numbers of larvae in the pasture, which reduces the reinfection rate among the animals and leaves them capable of developing natural immunity against nematodes.
Multidrug resistance parasites against imidothiazo- les (Sangster 1999), benzimidazoles (Silvestre & Cabaret 2002), and moxidectin (Blackhall et al., 1998), have been described in Brazil (Thomaz–Soccol et al. 2004). The spre- ad of resistant parasites is also increased by the intensive interregional transit ofsheep and goats. Sheep breeders of MG have been purchasing animals from the Northeast of Brazil, allowing the introduction of unkonwn parasites background. The limited use of quarantine for incoming animals in MG (Table 1) can exacerbate the situation allo- wing the introduction of different infectious agents such as helminths. According to Coles and Roush (1992), from the United Kingdom, the likelihood of spreading anthelmintic- -resistant nematodes could increase with the purchase of animals from other regions. In state of São Paulo, 81.3% of the sheep breeders used quarantine (Niciura et al. 2010) showing a significant difference regarding their level of in- formation when compared to the present study.
Gastrointestinalnematodes (GIN) impair animal productivity through reduction in voluntary food intake and/or inefficient use of nutrients. Disturbance in protein metabolism and reduced absorption and/or retention of minerals are significant during parasite infection (Waller, 1987). The controlof GINs predominantly depends upon chemotherapy despite advancements in genetical, immunological and biotechnological methods (Lloyd- Evans, 1991). Development of drug resistance in the parasites, effect of drug residues on human (Bowman et al., 1999) and high costs of the synthetic drugs, however, have led to attention of the workers to find the alternatives to the existing chemicals used for the treatment and/or controlof parasites (Al-Shaibani et al., 2009; Bachaya et al., 2009; Deeba et al., 2009; Sindhu et al., 2010). Plants
Braga et al. (2009) reported that it was unknown whether using these species in combination might add some kind of advantage from a biological point of view. Furthermore, according to Mota et al. (2003) , it would be important to assess the action of these fungi under in vivo conditions, irrespective of whether adminis- tered separately or in combination, so that their action could be observed in the fecal environment. In this regard, Tavela et al. (2013) observed the viability of coadministration of D. ﬂagrans and M. thaumasium on L3 cyathostomins after passage through the gastrointestinal tract of horses. However, subject to certain particularities, there have not been any reports showing that coad- ministration of these fungi on nematodes was capable of controlling these parasites in sheep in ﬁelds.
80% lower than those encountered in the paddock occupied by the control group were attained. The chemical group differed statistically from the control group only on day 30, and reached the end of the experiment with levels of pasture infestation due to L3 that were only 22.5% lower than those of the control group. Pasture irrigation may have favored fungal development. However, it may also have increased the challenge, as it also favors a better development ofnematodes. Dias et al. (2007) also observed reductions in the numbers of L3/kg of dry matter, of 85%, after six months of administering pellets of D. flagrans to cattle in southeaster Brazil. According to these authors, use of nematophagous fungi enabled reduction of the numbers of infective larvae in the pasture after one month. However, Larsen et al. (1998) only observed a statistical difference (p < 0.05) in the numbers of L3/kg of dry matter among horses treated with D. flagrans, in relation to the control group, in the last two months of their experiment.
The Fecal Egg Count Reduction Test (FECRT) was conducted to detect the presence of anthelmintic resistance. The flocks were divided in homogeneous groups of 10 to 20 animals with similar age, weight and sex. The number of groups was related to the number of treatments and their number of associations to be tested. One untreated group was used as control reference. Once the animals were weighed, their faeces were collected and the animals were treated and marked. All samples were kept in cold boxes until they were sent to the laboratory (Biology Sciences Dept., Veterinarian Parasitology of Universidade Federal do Paraná). A sample collection was repeated fourteen days after first treatment using identical procedures as first collection.
Parasite control is usually carried out with the use of anthelmintics, often indiscriminately and without any epidemiological knowledge (Lara 2013). The use of such suppressive regime added to inaccurate management strategies, contributes to the selection and establishment of parasites that are resistant to the anthelmintics (Almeida et al. 2010). As parasite resistance is an inherited trait, after every generation there may be an increase in individual parasites that would be able to survive drug treatment (Molento 2004). Parasite resistance is an alarming scenario in small ruminants worldwide, but there are no data on the magnitude of parasite resistance in sheep in Colombia. Therefore, the aim of this study was to report the lack of efﬁ cacy of the most used anthelmintics against gastrointestinalnematodesofsheep in Colombia.