Sheep and goat immune responses to nose bot infestation: a review.

Oestrus ovis L. (Diptera: Oestridae) is a cosmopolitan agent of myiasis in sheep and goats. The parasitic phase begins after adult females deposit first-stage larvae (L1) into the nostrils of hosts; these larvae develop into L2 and L3 in the nasal and sinus horn cavities. Sneezing and nasal discharges are the major clinical signs in infected animals. The pathogenesis of O. ovis infection is caused by: (a) the trauma resulting from the mechanical action of spines and hooks during larval movement on mucosal membranes, and, more importantly, (b) an allergenic reaction provoked by molecules excreted/secreted by larvae, of which salivary antigens are those mainly recognized by the host's immune system. The recruitment of immune reactive cells increases gradually from the nasal to sinus cavities in infected hosts. Mast cells, eosinophils, macrophages and lymphocytes are always more numerous in infected than non-infected animals. Humoral (antibody) systemic response of immunoglobulin G (IgG) usually reaches seroconversion 2-4 weeks post-first infection and the highest levels are observed during the development of L2 and L3 larvae. Local antibody responses include specific IgG, which has been found to negatively correlate with larval survival and development. Hypersensitivity reaction, immunomodulation, immunization trials and mixed infections of O. ovis and helminths are discussed.

IgG antibody response against salivary gland antigens from Oestrus ovis L. larvae (Diptera: Oestridae) in experimentally and naturally infected goats.

The aims of this study were to analyze the systemic IgG responses against third-instar salivary gland (L3SG) antigens by ELISA in Oestrus ovis experimentally infected kids (EIK) and in naturally exposed adult goats (NEG). Firstly, kids (n=4 per group) were assigned to receive intranasally 0, 12, 24, 36, and 48 first-instars in experimental infections. Blood samples were taken from EIK at Days 0, 14, 42 and 67 post-infection. At necropsy (Day 67), larval number and developmental instars were recorded. In an epidemiological study, blood serum samples were collected from 448 grazing NEG (n=20 flocks) in Baja California Sur, Mexico. Results showed that larval establishment rate was similar in EIK groups. Systemic IgG response reached the threshold after Day 42, but humoral response was not statistically different among EIK groups receiving experimental infections. In NEG, all surveyed flocks (100%) showed specific systemic IgG antibodies to L3SG antigens and the overall goat oestrosis prevalence was 59.2%. In conclusion, larval L3SG antigens were effective in detection of specific systemic IgG antibodies against O. ovis infected kids and goats by ELISA.

Specific IgG antibody responses in Oestrus ovis L. (Diptera: Oestridae) infected sheep: associations with intensity of infection and larval development.

Larvae of Oestrus ovis (Diptera: Oestridae) are ubiquitous parasites of nasal and sinusal cavities of sheep and goats. According to the chronobiology of O. ovis infections in Sardinia and the seasonal pattern of the IgG response, the optimal period to investigate the relationships between O. ovis larval populations and intensity of local and systemic IgG antibody responses was mid-July in the summer season. Sarda x Lacaune ewes (n=186), divided into three ram-families were used in the study. Systemic and local IgG responses were measured by ELISA tests using second stage larval crude extracts (L2CE) and L2 (L2SGC) and L3 (L3SGC) salivary gland contents as coating antigens. The number of larval instars, larval length of L1, L2 and L3 larvae, and larval weight of L2 and L3 larvae were individually recorded after ewe necropsy. Negative correlations among larval establishment and/or larval development on the one hand and intensity of local or systemic IgG responses on the other hand were found in two out of three studied ram-families.

Proteolytic activity in salivary gland products of sheep bot fly (Oestrus ovis) larvae.

This study identified and characterized hydrolytic enzymes in salivary gland products of Oestrus ovis larvae. Third instars were collected from the heads of slaughtered goats. Salivary glands were extracted, their products obtained by centrifugation and the enzymatic profile determined. Optimum pH, temperature of maximum proteolytic activity, thermal stability, and resistance of salivary gland products were determined on collagen and subclasses of proteases were identified using protease inhibitors. Zymograms were used to determine the molecular weight of proteases. Antigenic protein bands were revealed by immunoblotting using sera obtained from experimentally infested goats. Seven positive enzymatic activities were detected in salivary gland products: acid phosphatase, naphthol-AS-BI-phosphohydrolase, esterase (C4), esterase lipase (C8), leucine arylamidase, alpha-glucosidase and N-acetyl-beta-glucosaminidase. Optimum pH for proteolytic activity was 8.0; proteolytic activity increased with temperature (10-50 degrees C) then drastically decreased at 60 degrees C. Proteases in O. ovis salivary gland products belong to the serine subclass. In Zymograms, bands of proteolytic activity were detected in the 20-63 kDa range; the immunoblot showed three antigenic bands, one of them related to a protease band (63 kDa). Serine proteases in O. ovis salivary gland products are most likely involved in larval nutrition and host immuno-modulation.

Effects of immunization of Pelibuey lambs with Oestrus ovis digestive tract protein extracts on larval establishment and development.

Larval midgut proteins of hematophagous parasites contain strong antigens that can be used for host immunization. This concept has been applied for immunization of Pelibuey sheep against Oestrus ovis L. (Diptera: Oestridae). The aim of this study was to examine the effect of immunization on larval establishment (LE) and development. Immunized lambs (I, n = 6) received two injections of crude gut membrane protein extracts (GMPE) from third instar larvae with Freund's incomplete adjuvant (FIA) on days 0 (Day of first immunization) and 21 (0.4 and 0.45 mg GMPE/lamb, respectively). The control group (C, n = 5) received physiological saline with FIA. Lambs were challenged with first instars on Day 29 (20 larvae) and Day 43 (25 larvae). Blood samples were collected biweekly and IgG titers were analyzed by ELISA. All lambs were slaughtered on Day 90 and number of larvae recovered, larval stage and larval weight were recorded at necropsy. No significant effect of immunization on LE (C = 28.9%; I = 31.0% P > 0.05) was observed. Antibody titers were higher in the immunized group on Day 28 (P < 0.05), but subsequently similar in both groups. Larval physiological age and weight were also significantly (P < 0.05) affected by immunization. Immunization of Pelibuey lambs with GMPE did not affect LE but did delay O. ovis larval development.

In vitro and in vivo effects of neem tree (Azadirachta indica A. Juss) products on larvae of the sheep nose bot fly (Oestrus ovis L. Díptera: Oestridae).

Two studies were carried out in order to test the effects of neem tree extracts (Azadirachta indica A. Juss) on sheep bot fly larvae (Oestrus ovis L. Diptera: Oestridae). First, aqueous extracts from neem seeds (ASNE) at 0, 5 y 10% (w/v) concentrations were tested on larval mortality in vitro. In a second study, the effect of oral administration with neem seed meal (0, 100 y 200mg/kg) and neem leaves (1% of diet) on number of larvae found at necropsy and larval development was evaluated in experimentally O. ovis-infected sheep. Results in Experiment 1 showed a significant (P<0.05) effect of ASNE on time to L1 mortality in a dosis-dependent manner. In Experiment 2, oral administration of seeds or leaves did not affect the number of larvae found at necropsy of the sheep, but interfered with larval development and there was a tendency to reduce larval weight at the end of the infection period (55d).

Antioxidant enzymes in erythrocytes from goats seropositive to the sheep nose bot fly (Oestrus ovis L., Diptera: Oestridae) infection.

Oestrus ovis (Diptera: Oestridae) causes an important cosmopolitan parasitosis of the nasal and sinusal cavities of sheep and goats called oestrosis. Our objective was to analyze the participation of erythrocytes in the antioxidant system in goats seropositive to O. ovis infection under field conditions. Fifty female goats naturally exposed to O. ovis infection from Baja California Sur, México, were blood-sampled. Erythrocytic intracellular content was obtained from blood plasma. Oestrosis serodiagnosis was determined by ELISA. Protein, hemoglobin (Hb), superoxide dismutase (SOD), mieloperoxidase (MPO), catalase (CAT), glutathione-S-transferase (GST), and lipid peroxidation in erythrocytes were determined in both seropositive and seronegative goats. Overall seroprevalence of O. ovis infection in goats was 56%. Positive significant (P<0.05) associations were observed among systemic IgG level and protein (0.34), hemoglobin (0.43), SOD (0.32), and MPO (0.41) in erythrocytes. Protein and hemoglobin concentrations, as well as SOD and MPO activities in erythrocytes were found significantly higher (P<0.05) in seropositive than in seronegative goats. By contrast, enzymatic activities of CAT and GST and lipid peroxidation values were similar in seropositive and seronegative groups. In conclusion, there was a systemic stimulation of Reactive Oxygen Species which was efficiently scavenged by erythrocytic antioxidant enzymes in goats seropositive to O. ovis infection.