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1.
Acta Trop ; 201: 105213, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31600523

RESUMO

The aim of the present study was to evaluate the in vitro effect of quercetin combined with ivermectin (IVM) on Haemonchus contortus larvae and adults with different resistance profiles and demonstrate the in vivo anthelmintic action of this combination when used in sheep naturally infected. The effect of combination was evaluated based on the analysis of the mean effective concentration (EC50) obtained for larvae using the larval migration inhibition test and for adults using the motility test on females. The tests with larvae and adults were conducted using isolates with different degrees of susceptibility to IVM (sensitive, intermediate and highly resistant). The in vivo effect was evaluated based on the reduction in the egg count (FEC) and reduction in the count of adult helminths recovered after parasitological necropsy. Using the combination of quercetin with IVM, it was observed that in larvae, quercetin did not significantly reduce the EC50 for IVM in the sensitive and highly resistant isolates, but led to a significant reduction in the EC50 for IVM in the intermediate isolate. In adults, quercetin did not significantly reduce the EC50 for IVM in any of the isolates. No significant effect of the combination was found regarding the reduction in FEC or total count of parasites. The results of the in vitro and in vivo tests performed in the present study on quercetin activity underscore the importance of evaluating resistance-reversing agents among different stages of parasite development as well as among isolates with different resistance profiles. The action of quercetin combined with IVM on the motility of H. contortus larvae and adults was influenced by the degree of resistance and development stage of the parasite. The combination was effective only on intermediate resistant larvae. No action of the combination against adults was found. Moreover, this combination, when administered through the intra-abomasal route, was not effective at reducing the FEC and parasite load of naturally infected sheep.


Assuntos
Anti-Helmínticos/uso terapêutico , Resistência a Medicamentos/efeitos dos fármacos , Hemoncose/tratamento farmacológico , Haemonchus/efeitos dos fármacos , Ivermectina/uso terapêutico , Larva/efeitos dos fármacos , Quercetina/uso terapêutico , Doenças dos Ovinos/tratamento farmacológico , Animais , Feminino , Masculino , Contagem de Ovos de Parasitas , Ovinos/parasitologia , Doenças dos Ovinos/parasitologia
2.
Rev Bras Parasitol Vet ; 28(4): 644-651, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31800886

RESUMO

This study aimed to evaluate the anthelmintic activity of Eucalyptus citriodora essential oil and citronellal on sheep gastrointestinal nematodes. 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.


Assuntos
Anti-Helmínticos/farmacologia , Eucalyptus/química , Hemoncose/veterinária , Haemonchus/efeitos dos fármacos , Óleos Voláteis/farmacologia , Doenças dos Ovinos/parasitologia , Ovinos/parasitologia , Animais , Feminino , Cromatografia Gasosa-Espectrometria de Massas , Hemoncose/parasitologia , Haemonchus/isolamento & purificação , Haemonchus/ultraestrutura , Camundongos , Microscopia Eletrônica de Transmissão , Óleos Voláteis/isolamento & purificação , Contagem de Ovos de Parasitas , Testes de Sensibilidade Parasitária
3.
Exp Parasitol ; 207: 107780, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31629699

RESUMO

Plant extracts used for the treatment of helminth infections in sheep are an alternative to chemical anthelmintic drugs. Previous studies have reported the anthelmintic activity of acetone leaf extracts of Leucosidea sericea. For this study, we evaluate the ultrastructure changes induced by the acetone leaf extract of L. sericea and the component agrimol G (AG) that was isolated for the first time on adult haemonchus parasites. Adult haemonchus parasites harvested from sheep were incubated with the plant extract and AG for 3 h and evaluated by both scanning and transmission electron microscopy in comparison and in combination with albendazole or ivermectin. In all cases the method of evaluation shows ultrastructural changes, with albendazole inducing mitochondrial damage and ivermectin inducing muscle degeneration, both as previously described. Incubation with the plant extract and AG resulted in the formation of numerous non-membrane bound multi-vesicular like bodies and evenly spread disruptions/erosion in the epicuticle. Combining AG with ivermectin or albendazole resulted in an absence of effect of AG. Based on the structural changes induced by AG, together with the absence of an effect in combination with ivermectin and albendazole would suggest a disrupted microtubular network. The latter does however require biochemical confirmation.


Assuntos
Anti-Helmínticos/farmacologia , Haemonchus/efeitos dos fármacos , Floroglucinol/química , Extratos Vegetais/farmacologia , Rosaceae/química , Abomaso/parasitologia , Albendazol/farmacologia , Animais , Quimioterapia Combinada , Fixadores , Glutaral , Hemoncose/parasitologia , Hemoncose/veterinária , Haemonchus/ultraestrutura , Ivermectina/farmacologia , Microscopia Eletrônica de Varredura/veterinária , Microscopia Eletrônica de Transmissão/veterinária , Extratos Vegetais/química , Ovinos , Doenças dos Ovinos/parasitologia
4.
Nat Commun ; 10(1): 4811, 2019 10 22.
Artigo em Inglês | MEDLINE | ID: mdl-31641125

RESUMO

Haemonchus contortus is a haematophagous parasitic nematode of veterinary interest. We have performed a survey of its genome-wide diversity using single-worm whole genome sequencing of 223 individuals sampled from 19 isolates spanning five continents. We find an African origin for the species, together with evidence for parasites spreading during the transatlantic slave trade and colonisation of Australia. Strong selective sweeps surrounding the ß-tubulin locus, a target of benzimidazole anthelmintic drug, are identified in independent populations. These sweeps are further supported by signals of diversifying selection enriched in genes involved in response to drugs and other anthelmintic-associated biological functions. We also identify some candidate genes that may play a role in ivermectin resistance. Finally, genetic signatures of climate-driven adaptation are described, revealing a gene acting as an epigenetic regulator and components of the dauer pathway. These results begin to define genetic adaptation to climate in a parasitic nematode.


Assuntos
Anti-Helmínticos/farmacologia , Variação Genética , Haemonchus/efeitos dos fármacos , Haemonchus/genética , Animais , Clima , Resistência a Medicamentos , Genoma Helmíntico , Hemoncose/tratamento farmacológico , Hemoncose/parasitologia , Haemonchus/classificação , Haemonchus/isolamento & purificação , Humanos , Filogenia
5.
Parasit Vectors ; 12(1): 403, 2019 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-31412938

RESUMO

BACKGROUND: Haemonchus contortus, a gastrointestinal nematode parasite of sheep, is mainly controlled by anthelmintics; the occurrence of anthelmintic resistance leads to treatment failures and increases economic burden. Because molecular mechanisms involved in drug resistance can be elucidated by genomic studies, an extreme quantitative trait locus (X-QTL) mapping approach was used to identify co-segregation of the resistance phenotype with genetic markers to detect the genome-wide variants associated with monepantel resistance in H. contortus. METHODS: A cross between H. contortus isolates using parental susceptible (Par-S) males and monepantel resistant (Par-R) females resulted in SR progeny, while reciprocal cross resulted in RS progeny. Pools (n = 30,000) of infective larvae (L3) recovered from Par-R, and from SR and RS populations in the F3 generation, collected both before (unselected group) and 7 days after (selected group) selection with monepantel treatment in sheep hosts, were subjected to genome sequencing (Pool-Seq). Pairwise comparisons of allele frequencies between unselected and selected groups were performed for each population by Fisher's exact test (FET) and for both populations combined by a Cochran-Mantel-Haenszel (CMH) test. RESULTS: Mapping rates varied from 80.29 to 81.77% at a 90.4X mean coverage of aligned reads. After correction for multiple testing, significant (P < 0.05) changes in allele frequencies were detected by FET for 6 and 57 single nucleotide polymorphisms (SNPs) in the SR and RS populations, respectively, and by the CMH test for 124 SNPs in both populations. The significant variants located on chromosome 2 generated a selection signal in a genomic region harboring the mptl-1, deg-3 and des-2 genes, previously reported as candidates for monepantel resistance. In addition, three new variants were identified in the mptl-1 gene. CONCLUSIONS: This study expands knowledge on genome-wide molecular events underlying H. contortus resistance to monepantel. The identification of a genome region harboring major genes previously associated with monepantel resistance supports the results of the employed X-QTL approach. In addition, a deletion in exon 11 of the mptl-1 gene should be further investigated as the putative causal mutation leading to monepantel resistance.


Assuntos
Aminoacetonitrila/análogos & derivados , Anti-Helmínticos/farmacologia , Resistência a Medicamentos/genética , Haemonchus/efeitos dos fármacos , Haemonchus/genética , Locos de Características Quantitativas , Aminoacetonitrila/farmacologia , Animais , Feminino , Variação Genética , Masculino , Mutação , Fenótipo
6.
BMC Vet Res ; 15(1): 302, 2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31429752

RESUMO

BACKGROUND: Parallel in vitro and in vivo experiments were designed to evaluate promising chemotherapeutic alternatives for controlling haemonchosis in ruminants. In vitro anthelmintic activities (egg hatch test - EHT; larval development test - LDT) of aqueous and methanolic herbal extracts Mix1 and Mix2 were investigated. The in vivo effects of dietary supplementation with Mix1 and Mix2 on the parasitological status, inflammatory response, antioxidant parameters and microbial community of the lambs infected experimentally with Haemonchus contortus were investigated. Lambs were divided into four groups for the in vivo study: uninfected control lambs (C), infected lambs (I), infected lambs supplemented with Mix1 (I + Mix1) and infected lambs supplemented with Mix2 (I + Mix2). The experimental period was 70 days. RESULTS: The number of eggs per gram (EPG) of feces was quantified 22, 30, 37, 44, 51, 58, 65 and 70 days post-infection, and mean abomasal worm counts were assessed 70 days post-infection. Quantitative analyses identified 57.3 and 22.2 mg/g phenolic acids, 41.5 and 29.5 mg/g flavonoids and 1.4 and 1.33 mg/g protoberberine-type alkaloids in Mix1 and Mix2, respectively. The methanolic extracts of the herbal mixtures in both in vitro tests had higher anthelmintic effects (P < 0.01) than the aqueous extracts, but the effects did not differ significantly between Mix1 and Mix2 (P > 0.05). I + Mix1 and I + Mix2 lowered mean EPGs between 44 and 70 d by 58.1 and 51.6%, respectively. The level of IgG antibodies against H. contortus increased significantly after infection in each infected group. CONCLUSION: These results represent the first monitoring of the in vitro anthelmintic effects of herbal mixtures on H. contortus. The in vivo experiment indicated that the anthelmintic effect was not sufficient for the elimination of parasites, but this herbal treatment may affect the host over a longer term, reducing the parasitic infection in the host.


Assuntos
Produtos Biológicos/farmacologia , Hemoncose/veterinária , Doenças dos Ovinos/prevenção & controle , Animais , Feminino , Hemoncose/prevenção & controle , Haemonchus/efeitos dos fármacos , Contagem de Ovos de Parasitas , Plantas Medicinais , Ovinos
7.
Exp Parasitol ; 205: 107734, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31394093

RESUMO

Parasitism by Haemonchus contortus is one of the main limiting factors in small ruminant production around the globe. Although several studies suggest the use of integrated management practices, these parasites have been controlled essentially with synthetic anthelmintic drugs. The resistance mechanism against the imidazothiazole derivative levamisole in Haemonchus contortus has not been fully described. Recently, resistance was associated with a 63bp deletion in the Hco-acr-8b gene that encodes a subunit for a nicotinic acetylcholine receptor. This study aimed to standardize a real time PCR (qPCR) protocol for levamisole resistance diagnosis in H. contortus populations based on this polymorphism and use it to characterize 23 field H. contortus populations obtained from different localities of Ceará State, Northeast Brazil. In addition, two populations of H. contortus were used as a standard of susceptibility and resistance, Inbred Strain Edinburgh (ISE) and Kokstad, respectively. Larval development tests (LDT) were performed on five field isolates and both EC50 and EC95 were estimated. LDT EC95 values provided a wider interval between susceptible and resistant populations than EC50 values (EC95 = 1.96-57.93 µM; EC50 = 0.05-0.39 µM), and were found to be more appropriate for differentiating them. Real time PCR results showed resistance allele frequencies ranged from 20.9 to 76.7%. Our results suggest that levamisole resistance may be present in field populations but it is not as widespread as benzimidazole resistance. This methodology may be useful to monitor levamisole resistance in field populations of H. contortus.


Assuntos
Antinematódeos/farmacologia , Resistência a Medicamentos/genética , Haemonchus/efeitos dos fármacos , Levamisol/farmacologia , Animais , Benzimidazóis/farmacologia , DNA de Helmintos/isolamento & purificação , Fezes/parasitologia , Frequência do Gene/genética , Hemoncose/tratamento farmacológico , Hemoncose/parasitologia , Hemoncose/veterinária , Haemonchus/genética , Haemonchus/crescimento & desenvolvimento , Larva/efeitos dos fármacos , Larva/crescimento & desenvolvimento , Junção Neuromuscular/efeitos dos fármacos , Junção Neuromuscular/metabolismo , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Receptores Colinérgicos/efeitos dos fármacos , Receptores Nicotínicos/efeitos dos fármacos , Alinhamento de Sequência/veterinária , Ovinos , Doenças dos Ovinos/tratamento farmacológico , Doenças dos Ovinos/parasitologia , Tetramizol/farmacologia
8.
Vet Parasitol ; 273: 24-31, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31442889

RESUMO

Haemonchus contortus, one of the most pathogenic of all small ruminant parasites, have developed resistance to all used anthelmintics. Detoxification enzymes, e.g. cytochromes P450 (CYPs) and efflux transporters P-glycoproteins (P-gps), which represent the main defense system against harmful xenobiotics, have been suggested to contribute to drug resistance development. The present study was designed to compare the constitutive expression of individual CYPs and P-gps in females and males of H. contortus adults and to follow up on the changes in expression of these genes in nematodes exposed to sub-lethal concentrations of ivermectin (IVM), which might occur during inaccurate treatment. The adults of inbred susceptible-Edinburgh strain (ISE, MHco3) of H. contortus were used for this purpose. The nematodes were incubated ex vivo with or without IVM (1, 10 and 100 nM) in culture medium for 4, 12 and 24 h. After incubation, total RNA was isolated and expression levels of individual CYPs and P-gps were analyzed using qPCR. Our results showed a great variability in the constitutive expression of individual CYPs and P-gps in H. contortus adults. The constitutive expression as well as the inducibility of CYPs and P-gps significantly differed in males and females. Contact of adult nematodes with sub-lethal IVM concentrations led to only minor changes in expression of CYPs, while expression of several P-gps, particularly pgp-9.2 in males and pgp-10, pgp-11 in females was increased significantly in IVM-exposed nematodes. In conclusion, inaccurate treatment of sheep with IVM might contribute to drug resistance development via increased expression of efflux transporters in H. contortus adults.


Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Sistema Enzimático do Citocromo P-450/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Haemonchus/efeitos dos fármacos , Ivermectina/farmacologia , Animais , Resistência a Medicamentos/genética , Feminino , Haemonchus/genética , Masculino
9.
J Helminthol ; 94: e77, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31455460

RESUMO

This study assessed the in vitro anthelmintic (AH) activity of methanol and acetone:water leaf extracts from Annona squamosa, A. muricata and A. reticulata against Haemonchus contortus eggs. The egg hatch test was used to determine the effective concentrations required to inhibit 50% of eggs hatching (EC50). The role of polyphenols on AH activity was measured through bioassays with and without polyvinylpolypyrrolidone (PVPP). Methanolic extracts mainly caused the death of eggs at the morula stage (ovicidal activity). Meanwhile, acetone:water extracts caused egg-hatching failure of developed larvae (larvae failing eclosion (LFE) activity). The lowest EC50 values against H. contortus eggs were observed for the methanolic extracts from A. reticulata and A. muricata (274.2 and 382.9 µg/ml, respectively). From the six extracts evaluated, the methanolic extracts of A. muricata, A. reticulata and A. squamosa showed the highest ovicidal activity, resulting in 98.9%, 92.8% and 95.1% egg mortality, respectively. When the methanolic extract of A. squamosa was incubated with PVPP, its AH activity increased. Similarly, when acetone:water extracts of A. muriata and A. reticulata were incubated with PVPP, their LFE activity increased. Alkaloids were only evident in methanolic extracts, irrespective of PVPP incubation. The presence of acetogenins was not observed. In conclusion, methanolic extracts obtained from leaves of A. muricata, A. reticulata and A. squamosa showed ovicidal activity affecting the morula of H. contortus eggs, with minor LFE activity. Meanwhile, acetone:water extracts showed mostly LFE activity, with a lower proportion of ovicidal activity.


Assuntos
Annona/química , Anti-Helmínticos/farmacologia , Haemonchus/efeitos dos fármacos , Óvulo/efeitos dos fármacos , Extratos Vegetais/farmacologia , Acetona/farmacologia , Animais , Annona/classificação , Haemonchus/fisiologia , Larva/efeitos dos fármacos , Metanol/farmacologia , Folhas de Planta/química , Polifenóis/farmacologia
10.
Parasit Vectors ; 12(1): 330, 2019 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-31266521

RESUMO

BACKGROUND: Endoparasites are considered a major health problem of South American camelids as shown in a recent survey among German and Austrian camelid owners. Although prophylactic and therapeutic measures such as application of anthelmintics are commonly used, treatment efficacy is usually not assessed. Owners have expressed significant concerns regarding the effect of antiparasitic therapy, so this study aimed to evaluate the outcome of anthelmintic treatment in German alpaca herds with different drugs. RESULTS: Overall, 617 samples from 538 clinically healthy alpacas > 1 year-old from 27 farms (n = 11-157 animals/herd) were examined. The most common parasites detected by flotation were Eimeria spp. (75.1%) followed by strongylids (55.0%), Nematodirus spp. (19.3%), cestodes (3.1%) and Trichuris (2.7%). After initial coproscopical examination by flotation and strongylid egg quantification by the McMaster technique, positive animals excreting at least 150 eggs per gram of faeces were included in a faecal egg count reduction test (FECRT) using fenbendazole (n = 71 samples), moxidectin (n = 71) or monepantel (n = 66). Pre-treatment larval cultures (n = 23 positive pooled farm samples) revealed Haemonchus (87% of the farms), Cooperia (43.5%), Trichostrongylus (21.7%), Ostertagia (13.0%), Nematodirus and Oesophagostomum (4.3% each). Fenbendazole treatment reduced egg excretion by 45%, moxidectin by 91% and monepantel by 96%. On the farm level, 13/18 farms that used fenbendazole, 6/6 farms that used moxidectin and 2/5 farms that used monepantel had individual FECR values < 90% (fenbendazole) or < 95% (moxidectin, monepantel). Haemonchus and Cooperia were overrepresented on the farms with reduced treatment efficacy. CONCLUSIONS: Gastrointestinal strongylids are common in German alpacas and fenbendazole in particular was not sufficiently effective to reduce strongylid egg excretion. Although the FECRT could not unambiguously determine anthelmintic resistance in the present study, the finding that small ruminant strongylids, especially Haemonchus, are common in alpacas indicates that determination of effective anthelmintic doses, monitoring of efficacy and adapted (selective) treatment regimens must be implemented as part of sustainable deworming practices in this species in accordance with recommendations for ruminants.


Assuntos
Anti-Helmínticos/administração & dosagem , Camelídeos Americanos/parasitologia , Helmintíase Animal/prevenção & controle , Enteropatias Parasitárias/veterinária , Infecções por Strongylida/veterinária , Estrongilídios/efeitos dos fármacos , Aminoacetonitrila/administração & dosagem , Aminoacetonitrila/análogos & derivados , Animais , Fezes/parasitologia , Feminino , Fenbendazol/administração & dosagem , Haemonchus/efeitos dos fármacos , Helmintíase Animal/parasitologia , Enteropatias Parasitárias/parasitologia , Enteropatias Parasitárias/prevenção & controle , Macrolídeos/administração & dosagem , Masculino , Contagem de Ovos de Parasitas/veterinária , Infecções por Strongylida/parasitologia , Infecções por Strongylida/prevenção & controle
11.
Microb Pathog ; 135: 103629, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31325571

RESUMO

Lectins have been studied in the past few years as an alternative to inhibit the development of pathogenic bacteria and gastrointestinal nematodes of small ruminants. The development of new antibacterial and anthelmintic compounds is necessary owing to the increase in drug resistance among important pathogens. Therefore, this study aimed to evaluate the capacity of a glucose/mannose-binding lectin from Parkia platycephala seeds (PPL) to inhibit the development of Haemonchus contortus and to modulate antibiotic activity against multi-resistant bacterial strains, thereby confirming its efficacy when used in combination with gentamicin. PPL at the concentration of 1.2 mg/mL did not show inhibitory activity on H. contortus in the egg hatch test or the exsheathment assay. However, it did show significant inhibition of H. contortus larval development with an IC50 of 0.31 mg/mL. The minimum inhibitory concentration (MIC) obtained for PPL against all tested bacterial strains was not clinically relevant (MIC ≥ 1024 µg/mL). However, when PPL was combined with gentamicin, a significant increase in antibiotic activity was observed against S. aureus and E.coli multi-resistant strains. The inhibition of hemagglutinating activity by gentamicin (MIC = 50 mM) revealed that it may be interacting with the carbohydrate-binding site of PPL. It is this interaction between the antibiotic and lectin carbohydrate-binding site that may be responsible for the enhanced activity of gentamicin against multi-resistant strains. It can be concluded that PPL showed selective anthelmintic effect, inhibiting the development of H. contortus larvae and that it increased the effect of the antibiotic gentamicin against multi-resistant bacterial strains, thus constituting a potential therapeutic resource against resistant bacterial strains and H. contortus.


Assuntos
Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Fabaceae/química , Haemonchus/efeitos dos fármacos , Haemonchus/crescimento & desenvolvimento , Lectinas/farmacologia , Extratos Vegetais/farmacologia , Animais , Anti-Helmínticos/farmacologia , Gentamicinas/farmacologia , Haemonchus/microbiologia , Larva/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Sementes/química , Staphylococcus aureus/efeitos dos fármacos
12.
Vet Parasitol ; 271: 31-37, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31303200

RESUMO

The recovery of fenbendazole efficacy against Haemonchus contortus was attempted in a sheep intensive production system, using a strategy of population replacement in which the initial absolute efficacy of fenbendazole was 0%. The strategy was based on managing the parasite populations in refugia. Firstly, the resistant parasite population was reduced by means of anthelmintic treatments with efficacious drugs (Phase I), then a new, susceptible population was introduced in summer by way of artificially infected lambs at weaning, which were left to graze on the experimental pasture for eleven months (Phase II). Lastly, the impact of the replacement strategy, in terms of benzimidazole efficacy, was measured (Phase III). Faecal egg counts from permanent lambs and worm burdens as a measure of pasture infectivity from tracer lambs were determined throughout the study. During Phase I, faecal egg counts diminished from a peak of 2968 (300-7740) epg to 0 epg at the end, while adult worm burdens of H. contortus were reduced from 2625 (800-5100) to 0, which showed that the treatment strategy used in Phase I was effective in reducing the resistant population. These parameters also showed that good levels of pasture contamination and infectivity were achieved in Phase II, as faecal egg counts of up to 7275 (3240-13080) epg and adult worm burdens of 500 (200-800) H. contortus were reached. The absolute benzimidazole efficacy on H. contortus estimated at 16 months post-population replacement (Phase III) was 97.58%. The results lead to the conclusion that the recovery of anthelmintic efficacy of fenbendazole against a resistant population of H. contortus may be achieved by means of a strategy based on management of refugia and a subsequent introduction of a susceptible population. This strategy might be translatable to other resistant nematode genera.


Assuntos
Criação de Animais Domésticos/métodos , Resistência a Medicamentos , Fenbendazol/farmacologia , Hemoncose/veterinária , Haemonchus/efeitos dos fármacos , Criação de Animais Domésticos/normas , Animais , Anti-Helmínticos/farmacologia , Anti-Helmínticos/uso terapêutico , Benzimidazóis/farmacologia , Benzimidazóis/uso terapêutico , Fenbendazol/uso terapêutico , Hemoncose/tratamento farmacológico , Hemoncose/prevenção & controle , Ovinos
13.
Genome Biol Evol ; 11(7): 1965-1970, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-31263885

RESUMO

Internal parasitic nematodes are a global animal health issue causing drastic losses in livestock. Here, we report a H. contortus representative draft genome to serve as a genetic resource to the scientific community and support future experimental research of molecular mechanisms in related parasites. A de novo hybrid assembly was generated from PCR-free whole genome sequence data, resulting in a chromosome-level assembly that is 465 Mb in size encoding 22,341 genes. The genome sequence presented here is consistent with the genome architecture of the existing Haemonchus species and is a valuable resource for future studies regarding population genetic structures of parasitic nematodes. Additionally, comparative pan-genomics with other species of economically important parasitic nematodes have revealed highly open genomes and strong collinearities within the phylum Nematoda.


Assuntos
Anti-Helmínticos/farmacologia , Genoma Helmíntico/genética , Haemonchus/efeitos dos fármacos , Haemonchus/genética , Animais , Sequência de Bases , Genômica , Nova Zelândia
14.
BMC Vet Res ; 15(1): 184, 2019 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-31164118

RESUMO

BACKGROUND: Since pastoralists in South Darfur, Sudan, had complained about lack of albendazole (ABZ) efficacy to control nematodes in goats, the frequency of infection with gastrointestinal helminths was studied before in vivo faecal egg count reduction tests (FECRT) were conducted using ABZ orally either at the dose recommended for sheep, 5 mg/kg body weight (bw) or at 10 mg/kg bw. Experiments included goats naturally infected with gastrointestinal nematodes or experimentally infected with local Haemonchus contortus isolates. Three study areas (Nyala, Beleil and Kass) were visited in autumn or winter. RESULTS: Out of 478 screened goats, 82.4% were infected with gastrointestinal helminths and 82% were shedding eggs of strongyle nematodes with 90% of the strongyle larvae representing Haemonchus spp. A FECRT using naturally infected goats (n = 225: 71 untreated, 104 and 50 treated with 5 and 10 mg ABZ/kg bw, respectively) detected reduced ABZ efficacy in Nyala and Kass. Paired and unpaired FECRT calculations detected reductions of 72-92% with samples taken at 8 days post treatment with 5 mg ABZ/kg bw and of 85-94% with 10 mg ABZ/kg bw. The FECRT based on day 14 post treatment samples showed reductions of 69-77% with 5 mg/kg and of 75-87% with 10 mg ABZ/kg bw. In Beleil, ABZ efficacy was 95%. In the egg hatch test EC50 values for Nyala and Kass ranged from 0.12-0.24 µg thiabendazole/ml, corresponding to benzimidazole resistant phenotypes. Only Haemonchus spp. larvae were present after treatments in coprocultures. When the efficacy was evaluated experimentally using isolates of H. contortus from Nyala and Kass, the 5 mg ABZ/kg dose revealed reductions of 76-78% on day 8 and of 62-70% on day 14 with the unpaired method. Using 10 mg ABZ/kg, the FECR was still only 77-82%. CONCLUSIONS: Both, in vivo and in vitro methods detected resistant H. contortus populations in goats from South Darfur State. The time point 14 days post treatment was more sensitive for detection of ABZ resistance than 8 days post treatment. This is the first report on the occurrence of anthelmintic resistance in Sudan confirming that anthelmintic resistance selection is occurring in African subsistence farming systems.


Assuntos
Albendazol/uso terapêutico , Antinematódeos/uso terapêutico , Doenças das Cabras/parasitologia , Hemoncose/epidemiologia , Haemonchus/efeitos dos fármacos , Enteropatias Parasitárias/veterinária , Animais , Resistência a Medicamentos , Feminino , Doenças das Cabras/tratamento farmacológico , Doenças das Cabras/epidemiologia , Cabras , Hemoncose/tratamento farmacológico , Hemoncose/parasitologia , Enteropatias Parasitárias/epidemiologia , Masculino , Contagem de Ovos de Parasitas/veterinária , Prevalência , Sudão/epidemiologia
15.
Acta Parasitol ; 64(3): 638-644, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31172353

RESUMO

INTRODUCTION: Gastrointestinal nematodes (GINs) are a major constraint to the survival and productivity of animals. In southern China, goats are the most important small domestic ruminants. METHODS: From May 2013 to May 2017, we conducted a longitudinal study of hircine GIN infections in Huangshantou Town, Gongan County, Hubei Province, China, using fecal egg counts. RESULTS: Our investigation revealed that the GINs of goats in Hubei Province have changed significantly. Over 90% of eggs detected in the first month of investigation, May 2013, belonged to the species Haemonchus contortus and Chabertia sp. There was no seasonal variation in positive rates (PRs) of GINs, but the mean eggs per gram (EPG) of GINs were higher between April and July than between September and November (P < 0.05). The gradual increase in the percentage of H. contortus eggs among all detected eggs during our research and the low cure rate of IVM mass treatment revealed the emergence of IVM resistance in H. contortus. After the implementation of an integrated GIN control strategy, which included two mass treatments (one in April/May with ABZ and another in September/October with IVM + ABZ), in 2016 and 2017, both the PRs and EPG of GINs were significantly reduced. CONCLUSION: The results presented here reveal that controlling GINs of small ruminants in small farms in southern China requires an integrated control strategy that should include monitoring of infection and anthelmintic resistance, and increased farmer education on the importance of using the appropriate drugs at the correct dose.


Assuntos
Anti-Helmínticos/farmacologia , Resistência a Medicamentos , Gastroenteropatias/veterinária , Doenças das Cabras/parasitologia , Hemoncose/veterinária , Haemonchus/efeitos dos fármacos , Ivermectina/farmacologia , Animais , China , Feminino , Gastroenteropatias/parasitologia , Trato Gastrointestinal/parasitologia , Cabras , Hemoncose/parasitologia , Haemonchus/fisiologia , Estudos Longitudinais , Masculino , Estações do Ano
16.
Vet Parasitol ; 270: 13-19, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31213236

RESUMO

Small ruminants browsing in tropical forests readily consume the foliage of Senegalia gaumeri. A S. gaumeri methanol:water extract was recently shown to have ovicidal activity against Haemonchus contortus eggs in vitro. In the present study, the fraction of a S. gaumeri methanol:water extract with ovicidal activity against H. contortus eggs and the metabolites potentially involved in this activity were identified. Bio-guided fractionation of the S. gaumeri methanol:water extract identified high ovicidal activity (80.29%, EC50 = 58.9 µg/mL) in the non-polar sub-fraction P1. Gas chromatography-mass spectrometry (GC-MS) identified several fatty acids: pentacosane (18.05%), heneicosane (18.05%), triacontane (30.94%), octacosane (18.05%), and hexanedioic acid bis-(2-ethylhexyl) ester (32.72%). Purification of the polar components of sub-fraction P1 led to the identification of p-coumaric acid as a major constituent. In egg hatch tests, 400 µg/mL p-coumaric acid resulted in an ovicidal effect of 8.7%, a larvae failing eclosion effect of 2.9%, and of the emerged larvae (88.4%), many were damaged. In conclusion, the low AH activity of p-coumaric acid against H. contortus eggs indicates that it is not solely responsible for the ovicidal activity of sub-fraction P1 but might act in synergy with other compounds in this fraction. However, p-coumaric acid showed potential anthelmintic effects against the larval stage of H. contortus.


Assuntos
Anti-Helmínticos/isolamento & purificação , Anti-Helmínticos/farmacologia , Fracionamento Químico/métodos , Fabaceae/química , Haemonchus/efeitos dos fármacos , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/farmacologia , Animais , Larva/efeitos dos fármacos , Óvulo/efeitos dos fármacos , Folhas de Planta/química
17.
Vet Parasitol ; 270: 25-30, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31213238

RESUMO

This study evaluated the in vitro anthelmintic (AH) activity of cinnamic acid and six analogues against eggs and larvae of Haemonchus contortus. Stock solutions of each compound (trans-cinnamic acid, p-coumaric acid, caffeic acid, trans-ferulic acid, trans-sinapic acid, 3,4-dimethoxycinnamic acid, and chlorogenic acid) were prepared in PBS:Tween-20 (1%) for use in the egg hatch test (EHT) and larval exsheathment inhibition test (LEIT) at different concentrations (25-400 µg/mL). The respective effective concentration 50% (EC50) values with 95% confidence intervals were estimated. Mixtures made of all cinnamic acid and its analogues as well as some selected individual compounds were also tested in the EHT. Only ferulic and chlorogenic acids showed AH activity in the EHT (EC50: 245.2 µg/mL (1.26 mM) and 520.8 µg/mL (1.47 mM), respectively) (P < 0.05). A higher EC50 (1628.10 µg/mL) of the mixture of cinnamic acid and its analogues was required to observe activity against eggs mostly blocking the larvae hatching. The analogues' mixtures tested were less active than ferulic or chlorogenic acid alone. The activity of ferulic and chlorogenic acids against eggs was associated with larvae failing to hatch, and the two compounds exhibited antagonistic effects when evaluated together. All standards had an EC50 lower than 0.42 mM in the LEIT. Caffeic acid had the best activity in the LEIT (EC50 0.04 mM), followed by ferulic acid (EC50 0.11 mM) (P < 0.05). There was no clear, definitive structure-activity relationship for these non-flavonoid polyphenols against eggs or larvae of H. contortus in vitro. This study is the first to directly evaluate cinnamic acid and its derivatives as active compounds against eggs and larvae of H. contortus.


Assuntos
Cinamatos/farmacologia , Haemonchus/efeitos dos fármacos , Animais , Anti-Helmínticos/farmacologia , Cinamatos/química , Concentração Inibidora 50 , Larva/efeitos dos fármacos , Óvulo/efeitos dos fármacos
18.
Vet Parasitol ; 270: 40-46, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31213240

RESUMO

Parasitic nematodes pose a major threat to livestock production worldwide. The blood-feeding parasite Haemonchus contortus is a key small-ruminant pathogen that causes anaemia, and thereby seriously impacts animal health and production. Control of this parasite relies largely upon broad-spectrum anthelmintics, but new drugs are urgently needed to combat the threat of widespread multidrug resistance. Repurposing drugs can accelerate the development pipeline by reducing costs and risks, and can be an effective way of quickly bringing new antiparasitic drugs to market. Diarylamidine compounds such as pentamidine and diminazene have been employed in the treatment of trypanosomiasis and leishmaniasis in both human and veterinary settings, but their activity against parasitic worms has not yet been reported. We screened a small panel of diarylamidine compounds against H. contortus to assess their potential to be repurposed as anthelmintic drugs. Pentamidine and diminazene inhibited H. contortus larval development at low micromolar concentrations (IC50 4.9 µM and 16.1 µM, respectively, in a drug-susceptible isolate) with no existing cross-resistance in two multidrug resistant isolates and a monepantel-resistant isolate. Combinations of pentamidine with commercial anthelmintics showed additive activity, with no significant synergism detected. Pentamidine and diminazene showed different life-stage patterns of activity; both were active against early stage larvae in development assays, but only diminazene was active against the infective L3 stage in migration assays. This suggests some differences in uptake of the two drugs across the nematode cuticle, or differences in the nature and expression patterns of their molecular targets. As pentamidine and diminazene have been reported to be potent inhibitors of mammalian acid-sensing ion channels (ASIC), we tested the activity of known ASIC inhibitors against H. contortus to probe whether these channels may represent potential anthelmintic targets in nematodes. Remarkably, the spider-venom peptide Hi1a, a potent inhibitor of ASIC1a, inhibited H. contortus larval development with an IC50 of 22.9 ± 1.9 µM. This study highlights the potential use of diarylamidines as anthelmintics, although their activity needs to be confirmed in vivo. In addition, our demonstration that ASIC inhibitors have anthelmintic activity raises the possibility that this family of ion channels may represent a novel anthelmintic target.


Assuntos
Anti-Helmínticos/farmacologia , Diminazena/farmacologia , Haemonchus/efeitos dos fármacos , Pentamidina/farmacologia , Animais , Antiprotozoários/farmacologia , Técnicas In Vitro , Concentração Inibidora 50
19.
Parasit Vectors ; 12(1): 191, 2019 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-31039802

RESUMO

BACKGROUND: Due to anthelmintic resistance problems, there is a need to discover and develop new drugs for the treatment and control of economically important and pathogenic nematodes of livestock animals. With this focus in mind, we screened 236 compounds from a library (called the 'Kurz-box') representing chemically diverse classes such as heterocyclic compounds (e.g. thiazoles, pyrroles, quinolines, pyrimidines, benzo[1,4]diazepines), hydoxamic acid-based metalloenzyme inhibitors, peptidomimetics (bis- and tris-pyrimidoneamides, alkoxyamides) and various intermediates on Haemonchus contortus, one of the most important parasitic nematodes of ruminants. METHODS: In the present study, we tested these compounds, and measured the inhibition of larval motility and development of exsheathed third-stage (xL3) and fourth-stage (L4) larvae of H. contortus using an optimised, whole-organism phenotypic screening assay. RESULTS: Of the 236 compounds, we identified two active compounds (called BLK127 and HBK4) that induced marked phenotypic changes in the worm in vitro. Compound BLK127 induced an 'eviscerated' phenotype in the xL3 stage and also inhibited L4 development. Compound HBK4 exerted a 'curved' phenotype in both xL3s and L4s. CONCLUSIONS: The findings from this study provide a basis for future work on the chemical optimisation of these compounds, on assessing the activity of optimised compounds on adult stages of H. contortus both in vitro and in vivo (in the host animal) and against other parasitic worms of veterinary and medical importance.


Assuntos
Anti-Helmínticos/farmacologia , Haemonchus/crescimento & desenvolvimento , Animais , Anti-Helmínticos/química , Avaliação Pré-Clínica de Medicamentos , Feminino , Haemonchus/efeitos dos fármacos , Concentração Inibidora 50 , Larva/efeitos dos fármacos , Larva/crescimento & desenvolvimento , Masculino , Fenótipo
20.
Parasitol Res ; 118(4): 1215-1223, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30848351

RESUMO

Gastrointestinal nematodes remain a major constraint on the health, welfare, and production of small ruminants. This study was conducted to evaluate three plant extracts (from Bridelia ferruginea, Combretum glutinosum, and Mitragyna inermis) as effective remedies against gastrointestinal parasites of small ruminants. Phytochemical screening was conducted on the plant leaves, and the potential anthelmintic properties of these plants were tested in vitro on Haemonchus contortus using the egg hatch, larval migration, and adult worm motility assays. The phytochemical screening of the leaves revealed the presence of several bioactive components in all the plants. The number of eggs that hatched was reduced in a concentration-dependent manner (p < 0.01) upon treatment with the methanol extract of B. ferruginea and the acetone extracts of C. glutinosum and M. inermis. The inhibitory effect of the acetone extract of B. ferruginea and the methanol extracts of C. glutinosum and M. inermis was not concentration-dependent (p > 0.05). There was a significant difference (p < 0.05) in the reduction in larval migration between the lowest concentrations (75 to 150 µg/mL) and the highest concentrations (300 to 1200 µg/mL) of plant extracts. The ability of plant extracts to affect the mobility of the adult worms was not concentration-dependent (p > 0.05); however, it was dependent on the time of incubation (p < 0.01). At the highest concentration (2400 µg/mL), all adult worms were motionless after 24 h of exposure, while at the lowest concentration (< 150 µg/mL), this occurred after 48 h of exposure. M. inermis and C. glutinosum extracts were more effective than B. ferruginea extracts (p < 0.05). Overall, these results suggest that these plants used by small-scale farmers possess antiparasitic properties useful for helminthiasis control. However, the effects of the plants remain to be confirmed via in vivo assays and toxicity tests in further studies.


Assuntos
Anti-Helmínticos/farmacologia , Combretum/química , Haemonchus/efeitos dos fármacos , Malpighiales/química , Mitragyna/química , Compostos Fitoquímicos/farmacologia , Extratos Vegetais/farmacologia , Animais , Trato Gastrointestinal/parasitologia , Larva/efeitos dos fármacos , Locomoção/efeitos dos fármacos , Folhas de Planta/química , Ruminantes/parasitologia
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