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1.
Parasite Immunol ; 42(1): e12679, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31630404

RESUMO

Host protective immunity to Haemonchus contortus (Hc) infection in parasite-resistant St. Croix (STC) sheep is initiated early and characterized by an influx of innate cells and robust interleukin-4 (IL-4) production, resulting in T-helper type 2 immune (Th2) responses. The purpose of these studies was to elucidate the source of early IL-4 production. Neutrophils were isolated from whole blood, and populations >98% purity were cultured with larval or adult antigen to access cytokine production. Interleukin-4 and IL-13 were measured in sample supernatant using an ovine-specific enzyme-linked immunosorbent assay (ELISA). Neutrophils exposed to HcLA peaked in IL-4 production at 30 minutes (STC, 3153.65 pg/mL and SUF, 4665.22 pg/mL). A similar trend was observed in IL-13 production by 6 hours (STC, 391.02 pg/mL and SUF, 419.6 pg/mL). Adult antigen stimulation resulted in low cytokine production when compared to HcLA stimulation (STC IL-4, 6.04 pg/mL and SUF, 8.05 pg/mL, respectively; STC IL-13, 10 pg/mL and 12.5 pg/mL; P < .001), and no breed differences were observed. Mixed immune cell assays revealed an ability of neutrophils to induce IL-4 production in peripheral blood mononuclear cell (PBMC). Taken together, these data implicate neutrophils as a potential effector cell responsible for Th2 initiation.


Assuntos
Antígenos de Helmintos/imunologia , Hemoncose/veterinária , Interleucina-13/imunologia , Interleucina-4/imunologia , Neutrófilos/imunologia , Doenças dos Ovinos/imunologia , Animais , Cruzamento , Ensaio de Imunoadsorção Enzimática , Hemoncose/imunologia , Hemoncose/parasitologia , Haemonchus/crescimento & desenvolvimento , Haemonchus/imunologia , Interleucina-13/biossíntese , Interleucina-4/biossíntese , Larva/imunologia , Leucócitos Mononucleares/imunologia , Ovinos , Doenças dos Ovinos/parasitologia , Carneiro Doméstico , Células Th2/imunologia
2.
Parasite Immunol ; 42(1): e12680, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31631347

RESUMO

Eosinophils are prominent effector cells in immune responses against gastrointestinal nematode infections in ruminants, but their in vivo role has been hard to establish in large animals. Interleukin-5 is a key cytokine in the induction and stimulation of anti-parasitic eosinophil responses. This study attempted to modulate the eosinophil response in sheep through vaccination with recombinant interleukin-5 (rIL-5) and determine the effect on subsequent Haemonchus contortus infection. Nematode-resistant Canaria Hair Breed (CHB) sheep vaccinated with rIL-5 in Quil-A adjuvant, had lower blood eosinophil counts and higher mean worm burdens than control sheep vaccinated with Quil-A adjuvant alone. In addition, adult worms in IL-5-vaccinated sheep were significantly longer with higher eggs in utero in female worms, supporting an active role of eosinophils against adult parasites in CHB sheep. These results confirm that eosinophils can play a direct role in effective control of H contortus infection in sheep and offer a new approach to study immune responses in ruminants.


Assuntos
Eosinófilos/imunologia , Hemoncose/veterinária , Haemonchus/imunologia , Doenças dos Ovinos/imunologia , Adjuvantes Imunológicos , Animais , Gastroenteropatias/parasitologia , Hemoncose/imunologia , Interleucina-5 , Masculino , Contagem de Ovos de Parasitas , Saponinas de Quilaia , Ovinos , Doenças dos Ovinos/parasitologia , Carneiro Doméstico , Vacinação
3.
BMC Vet Res ; 15(1): 333, 2019 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-31533718

RESUMO

BACKGROUND: Haemonchosis is one of the most economically important parasitic diseases affecting small ruminants all over the world. Chemotherapeutic control has several shortcomings (limited anthelmintic arsenal, frequent resistance) and is hardly affordable by many farm economies. A recombinant antigen (rHc23) was shown to induce significant protection in vaccination trials with single dose challenges and different adjuvants. RESULTS: Lambs were vaccinated with 100 µg rHc23/dose + bacterial immunostimulant (BI) (LPS from Escherichia coli + Propionibacterium acnes extract) (days - 2, 0, 7 and 14) and subjected to a trickle infection with two dosages [6x, 1000 infective larvae (L3) or 6x, 2000 L3]. Vaccinated lambs showed a significant antibody response against rHc23 and Haemonchus contortus soluble extract as assessed by ELISA and Western blot (WB). Fecal egg counts (epg) along the experiment of vaccinated and BI treated lambs were significantly reduced. All vaccinated animals showed total egg output and abomasal helminth burdens (median, average) lower than those from unvaccinated or BI-treated animals lambs although differences were not statistically significant. CONCLUSIONS: Vaccination with 100 µg rHc23/dose + BI against H.contortus trickle infections apparently induced lower epg values and helminth burdens at the end of the experiment. Intragroup individual variations did not allow to obtain conclusive results and more research is needed including adjuvants and larger groups of animals to validate the potential value of rHc23 as candidate to develop a recombinant vaccine for lambs haemonchosis.


Assuntos
Antígenos de Helmintos/imunologia , Hemoncose/veterinária , Doenças dos Ovinos/parasitologia , Abomaso/parasitologia , Adjuvantes Imunológicos/administração & dosagem , Animais , Feminino , Hemoncose/imunologia , Hemoncose/prevenção & controle , Haemonchus/imunologia , Contagem de Ovos de Parasitas/veterinária , Proteínas Recombinantes/imunologia , Ovinos , Doenças dos Ovinos/imunologia , Doenças dos Ovinos/prevenção & controle , Carneiro Doméstico , Vacinação/veterinária
4.
J Anim Sci ; 97(11): 4428-4444, 2019 Nov 04.
Artigo em Inglês | MEDLINE | ID: mdl-31541548

RESUMO

The aim of this study was to identify for the first time single nucleotide polymorphisms (SNPs) associated with Haemonchus contortus resistance in Florida Native sheep, using a targeted sequencing approach. One hundred and fifty-three lambs were evaluated in this study. At the start of the trial, phenotypic records for fecal egg count (FEC), FAMACHA score, body condition score (BCS), and weight were recorded and deworming of sheep with levamisole (18 mg/kg of body weight) was performed. Ten days post-deworming (baseline) and 28 d post-baseline, a full hematogram of each sheep was obtained and FEC, FAMACHA score, BCS, and weight were assessed. Average daily gain was calculated at the end of the trial. Out of 153 animals, 100 sheep were selected for genotyping using a targeted sequencing approach. Targeted sequencing panel included 100 candidate genes for immune response against H. contortus. SNPs were discarded if call rate <95% and minor allele frequency ≤0.05. A mixed model was used to analyze the response variables and included the identity by state matrix to control for population structure. A contemporary group (age, group, and sex) was included as fixed effect. Bonferroni correction was used to control for multiple testing. Eighteen SNPs on chromosomes 1, 2, 3, 4, 6, 7, 11, 15, 18, 20, 24, and 26 were significant for different traits. Our results suggest that loci related to Th17, Treg, and Th2 responses play an important role in the expression of resistant phenotypes. Several genes including ITGA4, MUC15, TLR3, PCDH7, CFI, CXCL10, TNF, CCL26, STAT3, GPX2, IL2RB, and STAT6 were identified as potential markers for resistance to natural H. contortus exposure. This is the first study that evaluates potential genetic markers for H. contortus resistance in Florida Native sheep.


Assuntos
Resistência à Doença/genética , Estudo de Associação Genômica Ampla/veterinária , Hemoncose/veterinária , Haemonchus/imunologia , Doenças dos Ovinos/imunologia , Animais , Peso Corporal , Fezes/parasitologia , Feminino , Florida , Técnicas de Genotipagem/veterinária , Hemoncose/imunologia , Hemoncose/parasitologia , Masculino , Contagem de Ovos de Parasitas/veterinária , Fenótipo , Ovinos , Doenças dos Ovinos/parasitologia , Linfócitos T Reguladores/imunologia
5.
Parasitol Res ; 118(8): 2383-2388, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31203449

RESUMO

In this study, we evaluated in two trials a protocol designed to protect hair sheep using Barbervax®, a vaccine containing Haemonchus contortus gut membrane glycoprotein antigens. Results indicated that naturally infected vaccinated sheep had significant egg count reductions (90.2 ± 4.03%) compared with controls, although blood parameters remained relatively unchanged probably because the level of challenge was low. Vaccination prevented the periparturient rise in egg shedding of ewes, as well as egg shedding in lambs (37.1%). In the second trial, sheep which were experimentally exposed to higher artificial challenge also showed an efficient response to the vaccine as confirmed by high antibody levels and reduced egg counts and worm burdens (87 ± 5.4% and 79%) respectively. Thus, we believe that the vaccine should be integrated with other management practices for meat hair sheep as it has the advantages of adequate efficacy, reducing anthelmintic utilization and avoiding milk and environmental contamination with chemical residues.


Assuntos
Hemoncose/veterinária , Haemonchus/imunologia , Doenças dos Ovinos/prevenção & controle , Vacinação/métodos , Vacinas/administração & dosagem , Animais , Anticorpos Anti-Helmínticos/imunologia , Feminino , Hemoncose/imunologia , Hemoncose/parasitologia , Hemoncose/prevenção & controle , Haemonchus/genética , Contagem de Ovos de Parasitas/veterinária , Ovinos , Doenças dos Ovinos/imunologia , Doenças dos Ovinos/parasitologia , Vacinas/imunologia
6.
Parasite Immunol ; 41(7): e12625, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30883834

RESUMO

Interleukin 2 (IL-2) is an important immune regulatory factor in the immune response of the host. However, little is known about the inhibitor of host IL-2 in Haemonchus contortus infection. In this study, we found that globin domain-containing protein (HCGB) and Protein Y75B8A.8 (HC8) from H contortus excretory and secretory products are two binding proteins of IL-2 in goats. The yeast two-hybrid screening further validated the positive interactions of IL-2 with HCGB and HC8. Meanwhile, we found that HC8 had inhibitory effects on IL-2-induced peripheral blood mononuclear cell (PBMC) proliferation, while HCGB did not. Furthermore, transcriptional analysis revealed that HC8 could block the IL-2-activated signalling pathway. Our results showed that HC8 was a functional inhibitor of goat IL-2.


Assuntos
Doenças das Cabras/imunologia , Hemoncose/imunologia , Haemonchus/imunologia , Proteínas de Helminto/imunologia , Interleucina-2/antagonistas & inibidores , Animais , Doenças das Cabras/parasitologia , Cabras , Interleucina-2/imunologia , Leucócitos Mononucleares/imunologia , Transdução de Sinais
7.
Vet Parasitol ; 267: 47-53, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30878085

RESUMO

Haemonchosis commonly occurs as chronic and subclinical infection in small ruminants, and understanding of immunological response against subclinical haemonchosis is of paramount importance for designing and implementing effective control strategies. The present study was designed to evaluate immunological response during subclinical haemonchosis, experimentally established in goats. Sixteen 5-6 month-old helminth naive kids were randomly allocated into one of two groups, infected and uninfected; the infected group being infected per os with 250 Haemonchus contortus larvae per kg body weight. Faecal, blood and serum samples were collected every third day up to 30 days post-infection (DPI), thereafter weekly up to 58 DPI to record changes in faecal egg count (FEC), haemoglobin (Hb), packed cell volume (PCV), peripheral eosinophil percentage and immunological parameters, such as macrophage cytokine interleukin-12 (IL-12), Th1 cytokine (IFN-γ), Th2 cytokines (IL-4, 13, 25, 33) and immunoglobulins (IgG and IgE). Pre-patent period of H. contortus in the present study was 18 days and eggs per gram (EPG) peaked on 30 DPI. The total reduction in body weight gain in the infected group was 26 g per day when compared with uninfected animals. Hb (7.35 ± 0.34 g/dL in infected animals compared with 9.76 ± 0.67 in control animals) and PCV levels (22 ± 1.54 g/dL in infected animals compared with 29.2 ± 1.27 in control animals) decreased significantly up to 44 DPI in infected group (P = 0.000). IL-4, IL-13, IL-33, IgG and IgE showed significant increase in infected animals at different periods. IFN-γ, IL-12 and IL-25 did not show any significant changes barring a steep rise of IFN-γ on 27 DPI. A positive correlation was observed between IgE and IL-4 in subclinical haemonchosis. Of particular note was that all the major cytokines, such as IFN-γ (P = 0.000), IL-4 (P = 0.000), IL-13 (P = 0.009), and both IgG (P = 0.000) and IgE (P = 0.003), were observed at the lowest concentration on 24 DPI. The effect of infection was found to be significant on cytokines with a strong interaction with time. Taken together, the data suggest that Th2 immune response is predominating in subclinical haemonchosis. The economic loss in term of body weight gain due to subclinical haemonchosis was considerable.


Assuntos
Anticorpos Anti-Helmínticos/sangue , Citocinas/sangue , Doenças das Cabras/imunologia , Cabras/parasitologia , Hemoncose/veterinária , Animais , Infecções Assintomáticas , Peso Corporal , Fezes/parasitologia , Feminino , Hemoncose/imunologia , Haemonchus , Hematócrito , Imunoglobulina E/sangue , Imunoglobulina G/sangue , Índia , Contagem de Ovos de Parasitas , Células Th2/imunologia
8.
Animal ; 13(9): 1788-1796, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30700340

RESUMO

Breeding for resistance against nematodes has become the need of the hour due to emergence of anthelmintic resistant strains of major pathogenic nematodes of economic importance and rising demand for chemical residue free food by consumers. ICAR-Central Sheep and Wool Research Institute Avikanagar (Rajasthan) has developed Haemonchus contortus resistant lines of sheep in Avikalin and Malpura breeds by harvesting benefits of over-dispersion in fecal egg counts (FEC) through executing a breeding program since year 2004. Aim of the present study was to assess the genetic parameters for nematode resistance in these lines and also to develop suitable criteria for selection targeting resistance as well as growth improvement in these two lines. The data on 1240 Avikalin and 2172 Malpura sheep generated over 13 years (2004-16) for FEC along with deep pedigree and growth records for live weight at 6 (6WT) and 12 month were used for study. Data were analyzed using Average Information Restricted Maximum Likelihood (AIREML) approach. Results revealed moderate heritability (h2) for pre-drench log transformed fecal egg count (LFEC) in Avikalin (0.21±0.06) and Malpura (0.18±0.04) sheep. The post-drench h2 for LFEC was low in Avikalin (0.04±0.03) and Malpura (0.11±0.03) sheep. Effective selection program can be carried out for further improving the resistance against H. contortus in both the breeds using pre-drench LFEC estimates. The genetic correlation between the pre-drench LFEC and growth traits was not in the desired direction. Existence of substantial genotype × environment (G×E) interaction was seen in Malpura sheep, where major shift in ranks of sheep based on pre-drench LFEC as that of post-drench LFEC was observed owing to genetic correlation of 0.65±0.15. The G×E was absent in Avikalin sheep. Unreliable genetic correlation between growth and LFEC does not warrant a multi trait selection index development and its utilization in breeding program. The independent selection for LFEC followed by corrected 6WT can precisely help in achieving the goal of improving growth in nematode resistant sheep.


Assuntos
Anti-Helmínticos/farmacologia , Resistência à Doença/genética , Hemoncose/veterinária , Nematoides/crescimento & desenvolvimento , Doenças dos Ovinos/imunologia , Animais , Cruzamento , Fezes/parasitologia , Feminino , Hemoncose/imunologia , Hemoncose/parasitologia , Haemonchus/crescimento & desenvolvimento , Masculino , Nematoides/efeitos dos fármacos , Contagem de Ovos de Parasitas/veterinária , Linhagem , Fenótipo , Ovinos , Doenças dos Ovinos/parasitologia
9.
Parasite Immunol ; 41(2): e12611, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30548600

RESUMO

Haemonchus contortus is an economically important parasite that survives the host defense system by modulating the immune response. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is secreted by the parasite and the host responds by producing anti-enzyme antibodies. The enzyme inhibits complement cascade, an arm of the innate immunity, by binding to complement C3. In this study, the C3 binding site and the antigenic region of the enzyme were identified by generating short recombinant fragments and deleting a defined region of the enzyme. Using these proteins in ligand overlay and plate binding assay, the C3 binding region of GAPDH was localized within the 38 residues represented by 77-114 amino acids whereas one of the antigenic regions was identified in between 77 and 171 amino acids. In addition, deletion of amino acids 77 to 171 from GAPDH (fragment AB) also showed weak immunogenicity but lacked C3 binding activity. Fragment D comprising 95 residues (77-171), had both the C3 binding activity as well as immunogenicity like the parent enzyme, also stimulated host peripheral blood mononuclear cells in vitro. This truncated GAPDH moiety was stable at refrigerated temperature for at least 12 weeks and appears as a promising new therapeutic tool considering its longer shelf life as compared to the parent protein.


Assuntos
Complemento C3/metabolismo , Gliceraldeído-3-Fosfato Desidrogenases/imunologia , Haemonchus/imunologia , Animais , Sítios de Ligação , Ativação do Complemento , Gliceraldeído-3-Fosfato Desidrogenases/química , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Doenças das Cabras/imunologia , Doenças das Cabras/parasitologia , Cabras , Hemoncose/imunologia , Hemoncose/veterinária , Humanos , Leucócitos Mononucleares/imunologia , Ligação Proteica
10.
J Helminthol ; 93(4): 447-452, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-29669605

RESUMO

Many important studies on resistance reversion, anthelmintic efficacy and, especially, new molecules with antiparasitic effects are performed in laboratories using gerbils (Meriones unguiculatus) as the experimental model. This study aimed to evaluate the use of corticosteroids (dexamethasone and methylprednisolone acetate) in gerbils experimentally infected with different doses of infective larvae (sheathed or exsheathed) of Haemonchus contortus. In the first experiment, 28 gerbils were divided into seven groups infected by 2-6 × 103 larvae, with or without immunosuppression using corticosteroids. In the second experiment, eight gerbils were divided into two groups infected by 2 × 103 sheathed or exsheathed larvae. For the third assay, seven immunosuppressed gerbils were infected with 2 × 103 sheathed larvae and were killed 15 days post infection (PI). The highest number of parasites was recovered from methylprednisolone-immunosuppressed animals. We observed red and white blood cell alterations and biochemical parameters in infected animals that had undergone immunosuppression with methylprednisolone. We highlight that in the first and second experiments a satisfactory number of worms was recovered using sheathed larvae and immunocompetent animals. When exsheathed larvae were used, the number of worms recovered was unsatisfactory. A considerable larval burden was recovered from immunosuppressed gerbils 15 days PI, and body weight did not influence establishment of larvae.


Assuntos
Gerbillinae/parasitologia , Hemoncose/parasitologia , Haemonchus/patogenicidade , Imunossupressores/administração & dosagem , Corticosteroides/administração & dosagem , Animais , Modelos Animais de Doenças , Resistência a Múltiplos Medicamentos , Feminino , Hemoncose/imunologia , Haemonchus/efeitos dos fármacos , Imunossupressão/métodos , Larva/efeitos dos fármacos , Larva/patogenicidade , Masculino
11.
Vet Parasitol ; 264: 52-57, 2018 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-30503092

RESUMO

Haemonchus contortus is an economic problem in sheep farms worldwide, mainly in the tropics and subtropics. A vaccine against haemonchosis, called Barbervax®, was evaluated in ewes under two nutritional status, naturally infected with gastrointestinal nematodes. Ewes were divided into four groups: Supplemented Diet - Vaccine; Supplemented Diet - No vaccine; Basal Diet - Vaccine and Basal Diet - No vaccine. Their lambs were divided in Vaccinated and No vaccine. Ewes were immunised six times starting about 1 month of pregnancy with the first three doses at 3 week intervals and the last three shots at 4 week intervals. Supplemented ewes had higher body weight, body score and packed cell volume compared with those fed a basal diet. Both groups of vaccinated ewes showed a similar response in circulating anti-vaccine antibodies but the vaccine had no discernible effect on either body weight, body score and packed cell volume. There was a marked group difference in the number of ewes that received precautionary treatments with anthelmintic. All 14 Basal Diet - No vaccine ewes required treatment. In contrast only 7 ewes, in the Supplemented Diet - Vaccine group required anthelmintic treatment. In the Basal Diet - Vaccine and in the Supplemented Diet - No Vaccine groups, 12 and 13 ewes needed anthelmintic treatment, respectively. Vaccinated lambs showed much higher antibody titres resulting in 80% less Haemonchus spp. egg counts comparing with no vaccine lambs. Taken together these results clearly suggest that in pregnant and lactating ewes a combined protective effect between vaccination and improved nutrition resulted in fewer precautionary anthelmintic treatments. Thus, it was possible to achieve a more sustainable level of control of the haemonchosis, less dependent on anthelmintic drugs.


Assuntos
Suplementos Nutricionais , Hemoncose/veterinária , Nutrientes/imunologia , Doenças dos Ovinos/imunologia , Doenças dos Ovinos/prevenção & controle , Vacinas/imunologia , Animais , Anticorpos Anti-Helmínticos/sangue , Feminino , Hemoncose/imunologia , Hemoncose/prevenção & controle , Haemonchus/imunologia , Nutrientes/administração & dosagem , Gravidez , Ovinos , Clima Tropical , Vacinas/administração & dosagem , Ganho de Peso/imunologia
12.
Parasite Immunol ; 40(12): e12597, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30325524

RESUMO

Reduced worm burden in St. Croix (STC) sheep during Haemonchus contortus (Hc) infection is predicated on rapid interleukin-4 (IL-4) signalling and T helper type 2 immune (Th2) response. The aim of these studies was to further elucidate differences in Th2 responses by STC and Suffolk (SUF) sheep. Ten days after challenge Hc infection, peripheral blood mononuclear cells (PBMC) were collected and cultured with larval (HcLA) and adult worm (HcWA) antigen. STC PBMC produced nearly twice as much IL-4 as SUF (823.57 pg/mL vs 454.28 pg/mL) at 6 hours of HcLA culture despite no difference in IL-4 gene expression and the IL-4 receptor (IL4Rα) was upregulated in STC PBMC but was undetectable in SUF. Expression of other Th2-type genes were increased in STC PBMC including IL13, IL5 and MRC1. IL-4 supplementation to HcLA culture was insufficient to achieve upregulation of Th2 genes in SUF PBMC. Production of IL-4 did not occur in SUF PBMC until 24 hours after culture with HcLA, and expression of IL4 in the abomasum was similarly delayed until 10 days after challenge infection, which was associated with significantly higher larval burden (530 vs 16). These data demonstrate delayed upregulation of Th2 genes in SUF sheep contributes to susceptibility to Hc.


Assuntos
Hemoncose/veterinária , Haemonchus/fisiologia , Interleucina-4/imunologia , Doenças dos Ovinos/imunologia , Abomaso/imunologia , Abomaso/parasitologia , Animais , Feminino , Hemoncose/imunologia , Hemoncose/parasitologia , Haemonchus/genética , Haemonchus/imunologia , Leucócitos Mononucleares/imunologia , Masculino , Receptores de Interleucina-4/genética , Receptores de Interleucina-4/imunologia , Ovinos , Doenças dos Ovinos/parasitologia , Células Th2/imunologia
13.
Parasite Immunol ; 40(9): e12572, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29992577

RESUMO

This study aimed to characterize neutrophil response to Haemonchus contortus (Hc) in vitro using cells from parasite-resistant St. Croix (STC) and parasite-susceptible Suffolk (SUF) sheep. Neutrophils from Hc-primed and naive STC and SUF sheep were incubated with Hc larval antigen (HcLA), Hc worm antigen (HcWA) or complete media (CM). After HcLA exposure, neutrophils from STC and SUF formed extracellular traps composed of DNA. Stimulation with HcLA induced a 35-fold increase in extracellular DNA compared to CM controls. However, extracellular DNA was not found when neutrophils were cultured with HcWA. The formation of neutrophil extracellular traps (NET) in response to HcLA yields a low percentage of necrotic cells indicating a form of vital NETosis. Neutrophils from primed and naïve STC bound Hc L3 greater (93% and 68%) than SUF (78% and 45%; P < 0.001). Furthermore, STC neutrophils significantly reduced larval ATP levels compared to SUF neutrophils (0.05 µmol/L vs 0.1 µmol/L ATP, P < 0.001). These data indicate that ovine neutrophils bind, form vital NET and reduce ATP to Hc L3 in a breed and infection status-dependent manner.


Assuntos
Hemoncose/veterinária , Haemonchus/fisiologia , Doenças dos Ovinos/parasitologia , Animais , Cruzamento , Armadilhas Extracelulares/imunologia , Armadilhas Extracelulares/parasitologia , Hemoncose/imunologia , Hemoncose/parasitologia , Haemonchus/genética , Haemonchus/imunologia , Larva/genética , Larva/imunologia , Larva/fisiologia , Neutrófilos/imunologia , Ovinos , Doenças dos Ovinos/imunologia , Carneiro Doméstico , Especificidade da Espécie
14.
Parasite Immunol ; 40(9): e12569, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29959770

RESUMO

The experimental protocol of immunization tested here confirms its protective effect against Haemonchus contortus in goats. This protection translated into a 65.5% mean reduction in adult worm burden after a homologous challenge, and a significant decrease (73.2%) in cumulative faecal egg counts (FECs). These parasitological findings were consistent with the levels of some biopathological parameters. Thus, the reduction in adult worms and FEC observed in immunized animals were associated with increased levels of packed cell volume as well as plasma proteins. This response seems to be related to an important increase in specific antibodies (in serum and gastric mucus) and eosinophilia in response to challenge. At the local level, a cellular response was also observed in which CD4+ lymphocytes and globule leucocytes played a predominant role. Finally, it should be noted that the study of immunolocalization of proteins used in the vaccination trial suggests that these antigens have an internal location (at intestinal and reproductive tissues) in the adult worm. This observation, in conjunction with the kinetics of specific antibody levels after the challenge, suggests that these antigens may be part of excretory/secretory (E/S) products.


Assuntos
Doenças das Cabras/imunologia , Doenças das Cabras/prevenção & controle , Hemoncose/veterinária , Haemonchus/imunologia , Proteínas de Helminto/imunologia , Animais , Anticorpos Anti-Helmínticos/sangue , Fezes/parasitologia , Feminino , Doenças das Cabras/parasitologia , Cabras/imunologia , Hemoncose/imunologia , Hemoncose/parasitologia , Hemoncose/prevenção & controle , Haemonchus/genética , Proteínas de Helminto/administração & dosagem , Proteínas de Helminto/genética , Intestinos/imunologia , Intestinos/parasitologia , Masculino , Contagem de Ovos de Parasitas , Vacinação/veterinária , Vacinas/administração & dosagem , Vacinas/genética , Vacinas/imunologia
15.
Parasite Immunol ; 40(6): e12534, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29719931

RESUMO

The effect of Ovar-DRA and Ovar-DRB1 genotypes on faecal egg count (FEC) was determined in sheep and goats infected with Haemonchus contortus. One hundred and forty-three sheep from 3 different breeds (St. Croix, Katahdin and Dorper) and 150 goats from three different breeds (Spanish, Boer and Kiko) were used. Parasitological (FEC), haematological (packed cell volume) and immunological (IgA, IgG and IgM) parameters were measured. Sheep populations showed a higher FEC and humoural response than goat breeds. Genotypes were determined by high-resolution melting assays and by conventional PCR. For Ovar-DRA, sheep and goats carrying the AA genotype showed significant lower FEC than AG and GG genotypes. The additive effect was found to be 115.35 less eggs per gram of faeces for the A allele for goats. For Ovar-DRB1, only in sheep, the GC genotype was associated with low FEC. The additive effect was 316.48 less eggs per gram of faeces for the G allele, and the dominance effect was 538.70 less eggs per gram of faeces. The results indicate that single nucleotide polymorphisms within Ovar-DRA and Ovar-DRB1 could be potential markers to be used in selection programmes for improving resistance to Haemonchus contortus infection.


Assuntos
Doenças das Cabras/parasitologia , Hemoncose/veterinária , Haemonchus/imunologia , Infecções por Nematoides/veterinária , Contagem de Ovos de Parasitas , Proteínas de Ligação a RNA/genética , Doenças dos Ovinos/parasitologia , Proteínas Carreadoras de Solutos/genética , Animais , Fezes , Feminino , Genótipo , Cabras/parasitologia , Hemoncose/imunologia , Hemoncose/parasitologia , Haemonchus/crescimento & desenvolvimento , Polimorfismo Genético/genética , Ovinos/parasitologia
16.
Vet Parasitol ; 254: 105-113, 2018 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-29656994

RESUMO

We have set up an ex vivo ovine abomasal model, which can mimic the multicellular process to explore the early steps in haemonchine nematode infection using RNA-seq technology. Ovine abomasal explants were collected for histological and transcriptional analysis and supernatants collected to quantitate lactate dehydrogenase (LDH) enzymes. Atotal of 233 were substantially induced genes between L4-inoculated and uninoculated-control tissues, respectively. However, a total of 14 were considerably down-regulated genes between the 51 aforementioned tissues. Fifteen pathways were annotated by Kyoto Encyclopedia of Genes, and Genomes pathway analysis accounted for the significant percentage in immediate response to larval-stage of H. contortus. Key genes upregulated in response to the addition of L4-inoculum of H. contortus were IL-6, IL-8, C1q, Atypical chemokine receptor-3, chemokine ligand-2, manganese superoxide dismutase, integrin alpha-7, -8, -9, integrin subunit beta-1, integrin subunit beta 6, intercellular adhesion molecule-1 and actin alpha-1. This study shows for the first time that galectin-1 is up-regulated in an ex vivo abomasal segment model exposed to L4-inoculum of H. contortus following 6 h of incubation. The abomasal segment model has been shown to be a suitable tool to study the haemonchine larval-stage effects on the ovine abomasal tissues prior to in vivo assessment.


Assuntos
Abomaso/imunologia , Hemoncose/veterinária , Imunidade Inata , Doenças dos Ovinos/imunologia , Transcriptoma/imunologia , Animais , Modelos Animais de Doenças , Regulação para Baixo , Hemoncose/imunologia , Hemoncose/parasitologia , Haemonchus/crescimento & desenvolvimento , Haemonchus/fisiologia , Proteínas de Helminto/genética , Proteínas de Helminto/imunologia , Larva/crescimento & desenvolvimento , Larva/fisiologia , Ovinos , Doenças dos Ovinos/parasitologia , Regulação para Cima
17.
Proc Biol Sci ; 285(1874)2018 03 14.
Artigo em Inglês | MEDLINE | ID: mdl-29540516

RESUMO

It is normal for hosts to be co-infected by parasites. Interactions among co-infecting species can have profound consequences, including changing parasite transmission dynamics, altering disease severity and confounding attempts at parasite control. Despite the importance of co-infection, there is currently no way to predict how different parasite species may interact with one another, nor the consequences of those interactions. Here, we demonstrate a method that enables such prediction by identifying two nematode parasite groups based on taxonomy and characteristics of the parasitological niche. From an understanding of the interactions between the two defined groups in one host system (wild rabbits), we predict how two different nematode species, from the same defined groups, will interact in co-infections in a different host system (sheep), and then we test this experimentally. We show that, as predicted, in co-infections, the blood-feeding nematode Haemonchus contortus suppresses aspects of the sheep immune response, thereby facilitating the establishment and/or survival of the nematode Trichostrongylus colubriformis; and that the T. colubriformis-induced immune response negatively affects H. contortus This work is, to our knowledge, the first to use empirical data from one host system to successfully predict the specific outcome of a different co-infection in a second host species. The study therefore takes the first step in defining a practical framework for predicting interspecific parasite interactions in other animal systems.


Assuntos
Coinfecção/imunologia , Hemoncose/veterinária , Interações Hospedeiro-Parasita , Imunidade Inata , Doenças dos Ovinos/imunologia , Tricostrongilose/veterinária , Animais , Coinfecção/parasitologia , Hemoncose/imunologia , Hemoncose/parasitologia , Haemonchus/imunologia , Coelhos , Ovinos , Doenças dos Ovinos/parasitologia , Tricostrongilose/imunologia , Tricostrongilose/parasitologia , Trichostrongylus/imunologia
18.
Vet Parasitol ; 252: 180-186, 2018 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-29559145

RESUMO

Haemonchosis in camelids remains a challenging disease to treat, and prevention has become increasingly problematic due to widespread anthelmintic resistance. Barbervax®is an adjuvanted vaccine containing natural H-11, H-gal-GP antigens obtained from Haemonchus contortus adults via a proprietary process and solubilized in Quil A. This vaccine is approved for use in Australia, after demonstrating its safety and efficacy in sheep and goats. There are no published studies evaluating Barbervax in other ruminants/pseudoruminants such as camelids which can be parasitized with H. contortus. The vaccine utilizes a mixture of the parasite gut mucosal membrane enzymes including H-gal-GP and H11, involved in digesting a blood meal from the host. This study monitored the safety profile of the Barbervax® vaccine in a group of adolescent alpacas. Although designed into the original study of vaccine efficacy, the experimental infection with viable H. contortus third stage larvae could not be completed due to lack of detectable significant variation of infection following experimental challenge. Twelve alpacas (158 + 15 days) were randomized to vaccination with Barbervax® or no treatment. Three doses of Barbervax® were administered at 3 week intervals and investigators involved in animal monitoring and sample collection were blinded to the groupings. Clinical pathologic parameters were evaluated 7 days before vaccination, and 1 and 2 months post-vaccination. Daily clinical observations were made and specific observations regarding the injection site and rectal temperatures were monitored in each alpaca twice daily for 1 week following vaccination. Fecal egg counts, packed cell volume, and total protein were monitored following challenge with 1500 H. contortus larvae on days 42, 46, and 50. An increase in rectal temperature for a duration of 2 days (range 2-4 days) was observed post-vaccination. Vaccinated alpacas were lethargic for 2-3 days following vaccination; however, they maintained an appetite and no visible or palpable injection site reactions were observed. Following the first vaccination, all animals maintained normal clinical pathologic parameters throughout the study period. The vaccinated animals did develop titers to the H. contortus antigen as measured by ELISA. In conclusion, the Barbervax® vaccine demonstrated safety in this small group of young, healthy alpacas, but additional studies are required to evaluate the efficacy of the vaccine under field conditions in protecting alpacas against infection with H. contortus.


Assuntos
Hemoncose/veterinária , Haemonchus/imunologia , Vacinação/veterinária , Vacinas/efeitos adversos , Vacinas/imunologia , Animais , Anticorpos Anti-Helmínticos/sangue , Camelídeos Americanos/imunologia , Ensaio de Imunoadsorção Enzimática , Fezes/parasitologia , Hemoncose/imunologia , Hemoncose/prevenção & controle , Contagem de Ovos de Parasitas/veterinária , Vacinas/administração & dosagem
19.
Parasit Vectors ; 10(1): 425, 2017 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-28923082

RESUMO

BACKGROUND: Haemonchosis is a disease of the small ruminant caused by a nematode parasite Haemonchus contortus, and it is most important and alarming challenges to the small ruminant's production. The infection of the H. contortus could cause high economic losses worldwide. H. contortus is a blood feeding parasite which penetrates into the abomasal mucosa to feed the blood of the host and causing the anemia and decreased total plasma protein. Modulation and suppression of the immune response of the host by nematode parasites have been reported extensively, and the cysteine protease inhibitor (cystatin) is identified as one of the major immunomodulators. METHODS: The recombinant protein of HCcyst-3 was expressed in a histidine-tagged fusion soluble form in Escherichia coli, and its inhibitory activity against cathepsin L, B, as well as papain, were identified by fluorogenic substrate analysis. Native HCcyst-3 protein was localized by an Immunohistochemical test. The immunomodulatory effects of HCcyst-3 on cytokine secretion, MHC molecule expression, NO production and phagocytosis were observed by co-incubation of rHCcyst-3 with goat monocytes. RESULTS: We cloned and produced recombinant cystatin protein from H. contortus (rHCcyst-3) and investigated its immunomodulatory effects on goat monocyte. The rHCcyst-3 protein is biologically functional as shown by its ability to inhibit the protease activity of cathepsin L, cathepsin B, and papain. The immunohistochemical test demonstrated that the native HCcyst-3 protein was predominantly localized at the body surface and internal surface of the worm's gut. We demonstrated that rHCcyst-3 could be distinguished by antisera from goat experimentally infected with H. contortus and could uptake by goat monocytes. The results showed that the engagement of rHCcyst-3 decreased the production of TNF-α, IL-1ß and IL-12p40. However, it significantly increased the secretion of IL-10 and TGF-ß1 in goat monocytes. After rHCcyst-3 exposure, the expression of MHC-II on goat monocytes was restricted. Moreover, rHCcyst-3 could upregulate LPS induced NO production of goat monocytes. Phagocytotic assay by FITC-dextran internalization showed that rHCcyst-3 inhibited the phagocytosis of goat monocytes. CONCLUSIONS: Our results suggested that the recombinant cystatin from H. contortus (rHCcyst-3) significantly modulated goat monocyte function in multiple aspects.


Assuntos
Cistatinas/farmacologia , Cabras/imunologia , Haemonchus/imunologia , Proteínas de Helminto/farmacologia , Fatores Imunológicos/farmacologia , Monócitos/imunologia , Animais , Catepsina L/antagonistas & inibidores , Clonagem Molecular , Cistatinas/genética , Cistatinas/metabolismo , Citocinas/metabolismo , Escherichia coli/genética , Hemoncose/imunologia , Hemoncose/parasitologia , Haemonchus/química , Proteínas de Helminto/química , Proteínas de Helminto/genética , Proteínas de Helminto/imunologia , Monócitos/efeitos dos fármacos , Óxido Nítrico/metabolismo , Fagocitose/efeitos dos fármacos , Proteínas Recombinantes/farmacologia
20.
Vet Parasitol ; 243: 100-104, 2017 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-28807275

RESUMO

In our previous proteomics study, we identified Miro domain-containing protein (Miro-1), an excretory and secretory product of the pole worm, Haemonchus contortus, binds to goat peripheral blood mononuclear cells (PBMCs) in vivo. However, our understanding of the role of Miro-1and its potential immune impact on goat PBMCs is limited. The aim of the present study was to evaluate the effects of Miro-1 on functions of goat PBMCs in vitro. Recombinant protein (rMiro-1) was expressed in a prokaryote and incubated with goat PBMCs. Western blot analysis showed that rMiro-1 is successfully recognized by goat sera infected with H. contortus. Immunofluorescence analysis using rat antibodies against rMiro-1 indicated that this protein binds to goat PBMCs in vitro. Treatment of goat PBMCs/monocytes with various concentrations of rMiro-1 resulted in the upregulation of IL-2, IL-4, and IL-17, which in turn promoted cell proliferation, migration, the release of NO in PBMCs, and enhancement of phagocytosis of monocytes. These findings suggested that rMiro-1 stimulates PBMCs activity.


Assuntos
Hemoncose/veterinária , Haemonchus/imunologia , Proteínas de Helminto/farmacologia , Leucócitos Mononucleares/imunologia , Animais , Western Blotting/veterinária , Células Cultivadas , Cabras , Hemoncose/imunologia , Hemoncose/parasitologia , Proteínas de Helminto/genética , Interleucinas/metabolismo , Leucócitos Mononucleares/efeitos dos fármacos , Monócitos/imunologia , Proteínas Recombinantes/farmacologia
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