Tick-borne pathogens in ticks from urban and suburban areas of north-western Spain: Importance of Ixodes frontalis harbouring zoonotic pathogens.

To identify the questing tick populations in urban and suburban areas from the city of Lugo (NW Spain), ticks were collected monthly by flagging. The presence of Borrelia spp., Rickettsia spp. and Anaplasma phagocytophilum also was determined by polymerase chain reaction (PCR) and sequence analysis. Overall, 342 questing ticks were collected; the tick abundance was higher in suburban (95.9%) than in urban areas (4.1%). Ixodes frontalis was the most abundant (86.5%); 88.5% were larvae, 11.1% nymphs and 0.3% adults. All development stages of I. ricinus (7.3%) and adults of Rhipicephalus sanguineus sensu lato (5.8%) and Dermacentor reticulatus (0.3%) were found. Rickettsia spp. (31.9%) was more prevalent than Borrelia spp. (2.7%); no ticks were positive to A. phagocytophilum. Six Rickettsia species were identified (R. slovaca, R. monacensis, R. massiliae, R. raoultii, R. sibirica subsp. mongolitimonae and R. aeschielmanii); Candidatus Rickettsia rioja and two novel Rickettsia species also were detected. In addition, Borrelia turdi (1.8%) and B. valaisiana (0.9%) were identified in Ixodes ticks. This is the first report of R. slovaca in R. sanguineus s.l. and of R. monacensis, R. raoultii, R. slovaca, R. sibirica subsp. mongolitimonae and Ca. R. rioja in I. frontalis. Since most of the pathogens detected are zoonotic, their presence in these areas may have implications for public health.

Traumatic myiasis by Wohlfahrtia magnifica in sheep flocks from southeastern Spain: prevalence and risk factors.

Wohlfahrtia magnifica (Diptera: Sarcophagidae) is an obligate myiasis-causing fly of livestock that is widely distributed throughout the Mediterranean basin. A total of 73 683 sheep from 122 flocks in Albacete Province (southeastern [SE] Spain) were examined to detect active traumatic myiasis. The influence of several individual and flock factors on the prevalence was also assessed. The overall flock prevalence of traumatic myiasis was 95.9%, with an individual prevalence of 7.1%. Wohlfahrtia magnifica was identified in all of the positive animals except one that was infested by Lucilia sericata (Diptera: Calliphoridae). Adults of both genders showed a higher risk of infestation than juveniles, whereas males of all ages displayed an increased risk of infestation than females of all ages; the existence of a lambing building on the farm, poor husbandry facilities and the location area were also identified as risk factors. Prevalence was highest in the summer, and the fly season lasted from March to November. Genitalia were the main location of infestation in males (94.4%) and females (76.3%). Our results reveal that traumatic myiasis by W. magnifica is widespread in southeastern Spain, hence preventive measures minimizing the attractiveness of sheep to gravid flies are necessary to avoid health and welfare problems and significant production losses in sheep flocks.

Major differences in the larval anatomy of the digestive and excretory systems of three Oestridae species revealed by micro-CT.

Oestrid flies (Diptera: Oestridae) do not feed during the adult stage, so they depend on an efficient assimilation and storage of nutrients during their parasitic larval stage. We describe the general morphology and provide volumetric data for the digestive and excretory organs of the three larval instars of the nasal bot fly Oestrus ovis L., using micro-computed tomography. The size of the digestive and excretory organs greatly increased across larval instars. In all instars, the two salivary glands were remarkably large and formed a 'glandular band' by coming together, but without lumina uniting, at their posterior ends. The distal region of the anterior Malpighian tubules was greatly enlarged and full of highly radio-opaque concretions. Moreover, the anatomy of O. ovis third-instar larva was compared to that of two species of, respectively, similar and different feeding habits: Cephenemyia stimulator (Clark) and Hypoderma actaeon Brauer. Whereas the general morphology and arrangement of the digestive and excretory systems of C. stimulator was similar to that of O. ovis, some differences were observed in H. actaeon: a swollen anterior region of the midgut, salivary glands shorter and not forming a 'band' and anterior Malpighian tubules narrowly uniform throughout their entire length.

Molecular detection and identification of piroplasms (Babesia spp. and Theileria spp.) and Anaplasma phagocytophilum in questing ticks from northwest Spain.

Anaplasma phagocytophilum and some piroplasm species are pathogens mainly transmitted by Ixodes ricinus. Considering that this tick species is predominant in north-western Spain, individual specimens (652 nymphs, 202 females and 202 males) and 23 larval pools were processed to determine the prevalence of these pathogens in questing I. ricinus from that region. Additionally, Dermacentor marginatus, Dermacentor reticulatus, Ixodes frontalis and Ixodes acuminatus were individually analysed. The groESL operon as well as the 16S rRNA and msp2 genes of Anaplasma were analysed. Similarly, piroplasms were identified at the 18S rRNA gene and the ITS1 of Babesia spp. and Theileria spp. Babesia venatorum (1.5%), A. phagocytophilum (0.7%), Babesia microti (0.3%) and Theileria sp. OT3 (0.2%) were detected in I. ricinus. A single I. frontalis (8.3%) tested positive to A. phagocytophilum. Although a low percentage of I. ricinus were infected with A. phagocytophilum and piroplasms, a potentially human pathogenic variant of A. phagocytophilum was detected, and both Babesia species found were zoonotic. Since the vector of Theileria sp. OT3 remains unknown, further investigations are needed to unravel the role of I. ricinus in the transmission of this piroplasm.

Temporal and spatial spread of Hypoderma actaeon infection in roe deer from peninsular Spain determined by an indirect enzyme-linked immunosorbent assay.

The host switching of Hypoderma actaeon (Diptera: Oestridae), a specific parasite of red deer (Cervus elaphus), towards roe deer (Capreolus capreolus) has been recently reported in Spain. To provide information about the temporal and spatial spreading of H. actaeon infection in roe deer, 244 serum samples from animals hunted in Spain between 2013 and 2018 were analysed by an indirect enzyme-linked immunosorbent assay. The overall seropositivity was 13.9%. Seropositivity was higher in continental (27.7%) and mountainous (12%) areas from central Spain, followed by southern-Mediterranean (11.2%) and northern-oceanic regions (3.5%). Differences were significant between central-continental and northern-oceanic regions (P = 0.003). No differences were found according to the sex and age of roe deer (P > 0.05). In 2013, all seropositive animals were concentrated in two distant areas in central and southern Spain, suggesting that the host switch could have occurred independently in both regions. Changes in the pattern of distribution of red deer and roe deer could have favoured the spreading of this myiasis towards roe deer, indicating that roe deer may become infested by H. actaeon in areas where both cervids coexist at high densities.

A tooth whitening and chemical abrasive protocol for the treatment of developmental enamel defects.

This clinical report describes a chemical abrasive protocol and whitening techniques to quantify modifications to the color and volume produced when eliminating white spots associated with developmental defects in enamel. Chemical (oxidative-erosive) abrasive treatment (whitening + 6.6% hydrochloric acid) resolves white spots up to a depth of 0.2 mm. At greater depths, infiltration with 16% hydrochloric acid followed by composite resin infiltration is recommended.

Oral rehabilitation with dental implants in patients with recessive dystrophic epidermolysis bullosa: A retrospective study with 2-15 years of follow-up.

BACKGROUND: Epidermolysis bullosa (EB) comprises a group of hereditary disorders characterized by mechanical fragility of the skin and mucous membranes, with the development of blisters and vesicles in response to minimum tissue friction. Recessive dystrophic epidermolysis bullosa (RDEB) with generalized involvement is the most common subtype in the oral cavity. The present study was carried out to investigate dental implant survival, peri-implant tissue condition, patient satisfaction, and the impact of treatment upon the quality of life of patients with RDEB rehabilitated with implants and full-arch implant-supported prostheses. MATERIAL AND METHODS: Thirteen patients with RDEB underwent dental implant treatment between September 2005 and December 2016. A retrospective study was made to analyze implant survival, peri-implant tissue health and patient satisfaction. RESULTS: A total of 80 implants were placed (42 in the maxilla and 38 in the mandible) in 13 patients between 20-52 years of age and diagnosed with RDEB. All the implants were rehabilitated on a deferred basis with 20 full-arch prostheses. Fifteen fixed prostheses and 5 implant-supported overdentures were placed. The implant survival rate was 97.5% after a mean follow-up of 7.5 years after prosthetic loading. Fifty percent of the implants showed mucositis at the time of evaluation. Probing depth was maintained at 1-3 mm in 96.2% of the implants, and bleeding upon probing was observed in 67.5% of the implants. There was a high prevalence of bacterial plaque (85%). CONCLUSIONS: The treatment of edentulous patients with RDEB by means of implants and implant-supported prostheses is predictable as evidenced by the high success rate, and improves patient self-esteem and quality of life.

Occurrence of Borrelia and Borreliella species in Ixodes ricinus collected from roe deer in northwestern Spain.

Ixodes ricinus, comprising the predominant tick species in Europe, can transmit important human pathogens, including Borreliella spp., the causal agent of Lyme borreliosis. One hundred and seventy five roe deer hunted in two areas (plateau and mountain) of Galicia (northwest Spain) were examined for the presence of ticks; all roe deer were infested by I. ricinus. Nymphs (n = 1000), males (n = 1449) and females (n = 1000) of I. ricinus were analysed in pools of up to 10 ticks to detect both Borreliella and Borrelia DNA. The average number of I. ricinus per roe deer was similar in both areas, regardless of the life stage; although the percentage of Borreliella and Borrelia positive pools was higher in ticks collected from roe deer hunted in the plateau area, no significant differences were detected. Sequence analysis at the flagellin gene allowed the identification of four Borreliella species (Borreliella afzelii, Borreliella garinii, Borreliella lusitaniae and Borreliella valaisiana) and Borrelia miyamotoi in adult males; only B. valaisiana and B. miyamotoi were detected in nymphs and all females were negative. All Borreliella and Borrelia species found in roe deer were previously identified in questing I. ricinus collected in the same study area, although the prevalence was lower in the present study. The analysis of male I. ricinus ticks collected from roe deer gives a good estimation of Borreliella diversity in questing ticks.

Assessment of the capacity of a pyrophosphate-based mouth rinse to inhibit the formation of supragingival dental calculus. A randomized double-blind placebo-controlled clinical trial.

BACKGROUND: This study aimed to analyze the efficacy of an anti-calculus mouth rinse and its possible adverse effects on the mucosa and teeth. MATERIAL AND METHODS: This randomized double-blind placebo-controlled clinical trial included 40 patients with treated and managed periodontal disease, all with a history of rapid calculus formation. Patients used a pyrophosphate-based test mouth rinse (B) or a placebo (A). A range of parameters were measured for: saliva (saliva flow, pH and chemical composition); calculus (Volpe-Manhold [V-M] index, weight, and volume); adverse effects on mucosa and teeth; and the patients' subjective perceptive of mouth rinse efficacy. RESULTS: the test mouth rinse B produced reductions in urea, uric acid, and phosphorous, calcium, saliva flow, and increases in pH. V-M index and calculus weight decreased after using the test mouth rinse. Calculus volume decreased with both mouth rinses. No changes to the mucosa or teeth were observed. Patients perceived that the test mouth rinse was more effective. CONCLUSIONS: The test/B and placebo mouth rinses both modified certain parameters in saliva composition, particularly reductions in urea, uric acid, and phosphorous. Calcium tended to increase after using the test-B mouth rinse. The results did not demonstrate the anticalculus efficacy of the pyrophosphate-based mouth rinse or positive effects on saliva flow or composition. This field requires further research, as no product has been developed that prevents calculus formation completely.

Comparative in vitro study of the accuracy of impression techniques for dental implants: Direct technique with an elastomeric impression material versus intraoral scanner.

BACKGROUND: The aim of this study was to compare a conventional technique (elastomeric impression material - EIM) and a digital technique (scanner digital model - SDM) on a six-analog master model (MM) to determine which was the most exact. MATERIAL AND METHODS: Twenty impressions were taken of a master model (EIM) and twenty scanned impressions (SDM) (True Definition). A coordinate measuring machine (CMM) was used to measure the distances between adjacent analogues (1-2, 2-3, 3-4, 4-5, 5-6), intermittently positioned analogues (1-4, 3-6) and the most distal (1-6). Reference values were established from the master model, which were compared with the two impression techniques. The significance level was established as 5% (p<0.05). RESULTS: The precision of each technique was compared with MM. For adjacent analogues (1-2), no significant differences were found between EIM-MM (p=0,146). For intermittently positioned analogues (1-4), SDM did not show significant differences with MM (p=0.255). For the distance between distal analogues (1-6), significant differences were found between both techniques and MM (p=0.001). CONCLUSIONS: In a clinical situation with < three implants, EIM is more exact than SDM, but in cases of four implants SDM is more exact. For rehabilitations (> four implants), neither technique can be considered accurate although error falls within the tolerance limits established in the literature (30-150µm).

Hypoderma actaeon: an emerging myiasis in roe deer (Capreolus capreolus).

Subcutaneous larvae of Hypoderma spp. (Diptera: Oestridae) were detected in the dorsal region in 10 roe deer, Capreolus capreolus (Artiodactyla: Cervidae), hunted in central Spain between January and March 2016. All larvae were found in the inner side of the hide during the skinning of the animals. The study of the morphological features of eight larvae of different stages collected from two animals allowed the identification of Hypoderma actaeon Brauer. The small size (4-5 mm) of some of the first instars suggests that the internal lifecycle of H. actaeon may be exclusively subcutaneous. This is the first confirmation of H. actaeon in roe deer; however, further studies to assess the spread of the parasite and to follow the evolution of this myiasis in roe deer are needed.

Modulatory effects of interferon-γ and interleukin-4 on cellular immune responses against Hypoderma lineatum antigens.

This study investigates the in vitro modulatory effects of interferon-γ (IFN-γ) and interleukin-4 (IL-4) on both proliferative bovine T cell responses and IL-10 production induced by different antigens [crude larval extract and the purified fractions hypodermin A, B and C (HyA, HyB, HyC)] obtained from first instars of Hypoderma lineatum (Diptera: Oestridae), alone or in the presence of the mitogen concanavalin A. Incubation with the different parasitic antigens resulted in significant inhibition of T cell proliferation and IL-10 production, which, in general, did not revert after the addition of IFN-γ and IL-4. In the absence of antigens, IL-4 induced significant inhibition of mitogen-induced T cell responses. Exogenous IFN-γ exhibited an inhibitory effect on cell proliferation in the presence of the purified fractions HyB and HyC. These in vitro data suggest that far from neutralizing the effects of larval antigens, the addition of IFN-γ potentiates their anti-proliferative activity; by contrast, IL-4 had no consistent effects on proliferative responses to Hypoderma. IL-4 provoked an increment of IL-10 levels in supernatants of HyB-stimulated cells. In conclusion, exogenous IFN-γ and IL-4 were unable to counteract the suppressor effects of H. lineatum antigens.

A novel time-saving multiplex PCR assay for detecting and discriminating the most common canine Babesia species in Europe.

In Europe, most cases of canine babesiosis are caused by Babesia canis, Babesia vogeli (large piroplasms) and Babesia vulpes (small piroplasm). Molecular diagnosis is recommended due to its high sensitivity. Species identification after sequencing allows applying a rapid and efficient treatment, leading to a better prognosis; however, it is expensive and time-consuming. Thus, the objective of the present study was to develop a time-saving multiplex polymerase chain reaction (PCR) for simultaneously detecting and discriminating between large and small forms without sequence analysis. A new multiplex PCR was designed and tested using blood samples from 79 dogs showing clinical signs compatible with babesiosis which were previously analysed using blood smears and molecular methods. Multiplex PCR successfully discriminated between both Babesia groups showing bands of 700 and 890 bp for B. canis/B. vogeli and B. vulpes, respectively. No significant differences in the results of both PCR were detected and a substantial agreement between protocols (κ = 0.64) was found. Our multiplex PCR represents a reliable tool for detecting infections by the major Babesia spp. in dogs from Europe. Since no sequence analysis is required for identifying the species involved, this PCR allows the rapid administration of an appropriate treatment, thus improving the survival rate of the infected animals. In addition, it will represent a helpful tool for unravelling the real prevalence and distribution of B. vulpes and its implication in clinical cases.

Digital image analysis workflows for evaluation of cell behavior and tumor microenvironment to aid therapeutic assessment in high-risk neuroblastoma.

Digital pathology and artificial intelligence are promising emerging tools in precision oncology as they provide more robust and reproducible analysis of histologic, morphologic and topologic characteristics of tumor cells and the surrounding microenvironment. This study aims to develop digital image analysis workflows for therapeutic assessment in preclinical in vivo models. For this purpose, we generated pipelines that enable automatic detection and quantification of vitronectin and αvß3 in heterotopic high-risk neuroblastoma xenografts, demonstrating that digital analysis workflows can be used to provide robust detection of vitronectin secretion and αvß3 expression by malignant neuroblasts and to evaluate the possibility of combining traditional chemotherapy (etoposide) with extracellular matrix-targeted therapies (cilengitide). Digital image analysis added evidence for the relevance of territorial vitronectin as a therapeutic target in neuroblastoma, since its expression is modified after treatment, with a mean percentage of 60.44% in combined therapy tumors vs 45.08% in control ones. In addition, the present study revealed the efficacy of cilengitide for reducing αvß3 expression, with a mean αvß3 positivity of 34.17% in cilengitide treated material vs 66.14% in control and with less tumor growth when combined with etoposide, with a final mean volume of 0.04 cm3 in combined therapy vs 1.45 cm3 in control. The results of this work highlight the importance of extracellular matrix-focused therapies in preclinical studies to improve therapeutic assessment for high-risk neuroblastoma patients.

Specificity of Stenurus (Metastrongyloidea: Pseudaliidae) infections in odontocetes stranded along the north-west Spanish coast.

Parasites extracted from the lungs and the pterygoid sinus complex of 6 species of odontocetes stranded along the north-west Spanish coast (Northeast Atlantic) between 2009 and 2019 were morphologically identified. The samples belonged to 14 specimens, including 3 harbour porpoises, Phocoena phocoena, 6 short-finned pilot whales, Globicephala macrorhynchus, 1 long-finned pilot whale, Globicephala melas, 1 Risso's dolphin, Grampus griseus, 1 striped dolphin, Stenella coeruleoalba and 2 bottlenose dolphins, Tursiops truncatus. All animals (14/14) were infected by nematodes of the genus Stenurus spp.; moreover, two of them presented a mixed lung nematode infection by Stenurus spp. and Halocercus spp., and another two a mixed infection by Stenurus spp. and the trematode Nasitrema spp. in the pterygoid sinuses. The morphological characterization of the Stenurus specimens revealed the existence of three different species: Stenurus minor, present in the pterygoid sinuses of harbour porpoises with a mean intensity of 43.0 ± 9.0; Stenurus globicephalae, in the pterygoid sinuses of pilot whales and the Risso's dolphin (370.3 ± 579.4); and Stenurus ovatus infecting bottlenose and striped dolphins' lungs (47.7 ± 76.5). This is the first citation of S. minor and S. ovatus in odontoceti from the Galician coast. Nematodes of the genus Stenurus are frequent in odontocetes stranded along the north-west Spanish coast. A clear host-parasite association was observed between S. minor and the Phocoenidae family, between S. globicephalae and the subfamily Globicephalinae and between S. ovatus and subfamily Delphininae. Different trophic position and niche segregation may lead to different patterns of specificity.

Use of a crude extract or purified antigen from first-instar cattle grubs, Hypoderma lineatum, for the detection of anti-Hypoderma antibodies in free-ranging cervids from southern Spain.

During the 2003-2005 hunting seasons, a total of 120 Cervidae, including 39 red deer (Cervus elaphus hispanicus) and 81 fallow deer (Dama dama), were examined for subcutaneous myiasis. Animals were shot from January to June in southern Spain. Specific antibodies against Hypodermatinae (Diptera: Oestridae) were detected by indirect enzyme-linked immunosorbent assay (iELISA) using a crude larval extract (CLE) and a purified antigen [hypodermin C (HC)] obtained from first instars of Hypoderma lineatum (De Villers) (Diptera: Oestridae). Hypoderma actaeon Brauer was the only species detected in this study, which represents the first confirmation of this species in fallow deer from Spain. The overall prevalence of animals presenting subcutaneous larvae (14.2%) was considerably lower than the prevalences determined by iELISA with CLE (43.3%) and HC (40.0%). Red deer showed a higher prevalence of Hypoderma than fallow deer. The concordance between larval examination during the hunting season and iELISA using both antigens was low, whereas the concordance between the CLE and HC ELISAs was good. Larval antigens obtained from H. lineatum constitute a good tool for the diagnosis of H. actaeon in Cervidae, especially when the hunting season does not coincide with the maximum presence of larvae on the back.

Efficacy of anthelminthic control programs against natural Muellerius capillaris infection in sheep in the north-west of Spain. Effect on blood gases and pH in venous blood samples.

The field efficacy of a single dose treatment against natural M. capillaris infection using different anthelmintic drugs, extensively employed in ovine parasite control programs in Galicia (N.W. Spain), and the effect of protostrongylid infection on ovine respiratory functions, were evaluated. Five groups (n = 5) of ewes were used in this study; animals were treated with injectable ivermectin 10.2 mg/kg), levamisole (7.5 mg/kg) and albendazole (5 mg/kg) and monitored at 0, 7, 21, 42 and 63 days posttreatment (d.p.t.) by enumeration of the lungworm larvae per gram of faeces (l.p.g.) and determination of gas tension and pH in venous blood using an i-Stat portable clinical analyzer. No statistical difference was found either in larval elimination between untreated and treated groups or in the reduction in larval counts in all of the treated groups. A significant increase in partial oxygen tension (pO2) and saturation (sO2) between day 0 and 7 p.t. was observed in all treated animals. These values decreased significantly at day 21 to previous levels. There were no statistical differences in blood gases between uninfected and treated groups. We can conclude that under Galician field conditions, parasitic control programs are not totally effective against M. capillaris infection.

Cytokine expression in bovine PBMC cultures stimulated with Hypoderma lineatum antigens.

Hypoderma antigens are involved in host inflammation and immune response, conditioning larvae survival. In this study, peripheral blood mononuclear cell (PBMC) cultures from Hypoderma sensitized and unsensitized cattle were performed to determine the effect of H. lineatum antigens and incubation time (18, 24, 48 h) on IFN-γ, TNF-α, IL-10 and IL-4 mRNA gene expression determined by RT-qPCR. TNF-α and IL-4 gene expression were higher in Hypoderma previously sensitized PBMCs, suggesting that a mixed Th1/Th2 response may play a significant role in host defence reactions against Hypoderma exhibited by previously infested cattle. Incubation time had a significant effect on IL-10 and TNF-α gene expression, which decreased over time. Regarding to H. lineatum antigens, the crude larval extract and the purified fraction hypodermin B (HB) produced a significant reduction of the mRNA expression levels of the proinflammatory cytokine, IFN-γ; moreover, the HB had a stimulating effect on the mRNA gene expression of the anti-inflammatory cytokine IL-10, demonstrating that the parasite would modulate the host defence mechanisms by avoiding harmful immune responses that would limit its survival into the host tissues.

Immunomodulatory effect of Hypoderma lineatum antigens: in vitro effect on bovine lymphocyte proliferation and cytokine production.

This study examines the immunomodulatory effect of a crude larval extract (CLE), obtained from first stage larvae (L1) of H. lineatum, and the purified fractions hypodermin A (HyA), HyB and HyC. Proliferative responses of peripheral blood mononuclear cells (PBMC) from uninfested and previously infested cattle and the production of the cytokines IL-10, IL-4 and IFN-gamma, in response to concanavalin A (Con A), were determined. The stimulation index of peripheral blood mononuclear cells from uninfested cattle was significantly lower than that from infested animals with the different antigens assayed. The HyA was the antigen that most inhibited the proliferative response, followed by the HyB, the HyC and the CLE. This hypodermin provoked an increase of IFN-gamma and a suppression of IL-10 production that would support a Th1-like cytokine response. The HyB reduced the production of IL-10 stimulated by the Con A in cultures from infested animals. The HyC did not modulate the production of cytokines. Finally, the CLE induced a marked suppression in the production of the different cytokines in cultures from naïve and previously sensitized animals. Our results indicate that Hypoderma larval secretions are comprised of different components (hypodermins) that individually induce distinct but partially overlapping modulatory responses.

Impact of previous infestation on dynamics of circulating hypodermin C in cattle artificially infested with Hypoderma lineatum (Diptera: Oestridae).

Four groups of cattle were artificially infested with 50 first instar Hypoderma lineatum after either a primary natural infestation or in the absence of a primary infestation. In two groups the primary infestation had been terminated by the application of either an organophosphate insecticide or a macrocyclic lactone parasiticide. Circulating hypodermin C and specific antibodies were measured for 40 weeks after the artificial infestation. Stage specific mortality of the larvae was also monitored. Previously uninfested cattle exhibited typical antibody and antigen profiles during the infestations. Antibodies were first detected on Week 7p.i., they rose to maximum values between Weeks 24 and 25, then declined as larvae reached the back. A second peak occurred at Week 32 and antibody remained more less constant thereafter. Previously infested groups exhibited a dramatic anamnestic response by Week 3p.i. with antibody levels peaking at Week 8. A second peak was noted between Weeks 24 and 26p.i. after which antibody levels declined and then remained relatively stable. The dynamics of circulating hypodermin C in the previously infested cattle resembled those in the previously uninfested cattle. Mortality of first instars did not differ among the four groups. Similarly mortality of second and third instars, in the warble, did not differ although there was a tendency for higher mortality in the previously infested, untreated group. These results reinforce previous work demonstrating the development of a significant immune response during the primary infestation that is reflected in the rapid and substantial production of antibodies upon re-infestation. It is significant that a challenge model using subcutaneous injection of newly hatched first instars avoids host immune responses in the skin that result in substantial larval mortality. The current data also support the concept that migrating first instars induce significant reduction in host immune response. There is a peak of antibody production as antigen levels peak during first instar migration to the back. As larvae molt to the second instar and antigen production ceases there is a persistence of antibody which suggests release of the immune response from the suppression induced by the first instar secretory antigens.