Expression Patterns of Host Inflammatory Cytokine Genes during Infestation with Haemaphysalis longicornis, a Zoonotic Vector, in Blood Sucking Periods.

Tick saliva is critically important for continuous attachment to the host, blood feeding for days, and transmission of tick-borne pathogens. To characterize the patterns of inflammatory cytokine gene expression during its attachment and blood sucking time, peripheral blood samples of rabbits infested with Haemaphysalis longicornis ticks were collected at different intervals. Blood histamine concentration was evaluated as well as gene encoding IFN-γ, TNF-α, IL-2, IL-6, IL-4, and IL-10 were compared with non-infested rabbits. Blood histamine concentration of tick-infested rabbits during fast feeding time was significantly higher than that of non-infested rabbits. In both nymph and adult tick infested rabbits, expression of TNF-α and IFN-γ genes were decreased significantly (P<0.05), while expression of IL-4, IL-6, and IL-10 were increased 1.3 to 7 folds in adult infested rabbits with the exception of IL-6 that was significantly (P<0.05) decreased in nymph infested rabbits. IL-2 was not expressed in either nymph or adult infestation. H. longicornis saliva is capable of modulate host responses through a complex correlation with histamine and Th1, Th2 mediated cytokines that suppress the inflammatory responses directed toward inflammatory mediators introduced into the host during tick feeding.

The effect of tick infestation on the serum concentrations of the cardiac biomarker troponin I, acid-base balance and haematobiochemical profiles in camels (Camelus dromedarius).

The aim of the present study was to evaluate the serum concentration of cardiac troponin I (cTnI) in camels with tick infestation as a marker of potential myocardial injury and its prognostication. The effects of acid-base and electrolyte balance and haematobiochemical profiles were also investigated. Twenty-three camels (Camelus dromedarius) with tick infestation and suffering from anorexia, incoordination of movement, unsteady gait, recumbency, opisthotonus, anaemia and reduced production were examined. Ticks were visible, deeply embedded in the skin over the whole body, especially under the neck and around the udder or testis. Of the camels, 15 recovered after treatment and 8 did not. Blood samples were collected from the diseased camels on the day of admission to our clinic. Blood samples were also collected from 12 healthy camels and were used as controls. The mean serum concentration of cTnI in the camels with tick infestation was 1.7 ± 1.6 ng/ml compared to 0.03 ± 0.02 ng/ml in the controls. The mean serum concentration of cTnI in those camels that recovered was 0.36 ± 0.53 ng/ml compared to 3.0 ± 1.1 ng/ml in the camels that did not. Venous blood gas alterations included an increase in partial pressure of carbon dioxide and decreases in partial pressure of oxygen and oxygen saturation. Haematological parameters involved decreases of red blood cells, haemoglobin and haematocrit and increases in the mean corpuscular volume, mean corpuscular haemoglobin and mean corpuscular haemoglobin concentration. Biochemical parameters included significant elevations in the serum activity of aspartate aminotransferase, creatine kinase and alkaline phosphates. Other biochemical alterations observed were decreases in total protein and albumin and increases in globulin and glucose concentrations. In conclusion, the data of this study suggest the possibility of using cTnI as a biomarker for cardiac injury in camels with tick infestation and for the prognosis of the outcome in the treated animals. Generally, values above 1.0 ng/ml were considered bad prognostic indicators.

Hematological and plasma profiles and ticks and tick-borne pathogens in wild Formosan black bears (Ursus thibetanus formosanus).

BACKGROUND: The endangered Formosan black bear (Ursus thibetanus formosanus) is the largest native carnivorous mammal in Taiwan. Diseases, poor management, illegal hunting, and habitat destruction are serious threats to the survival of bear populations. However, studies on the impact of diseases on bear populations are limited. Therefore, this study aimed to establish a database of the hematological and plasma profiles of free-ranging Formosan black bears and investigate the occurrence of ectoparasites, blood parasites, and vector-borne pathogens. METHODS: Formosan black bears were captured in Yushan National Park (YNP) and Daxueshan Forest Recreation Area (DSY) in Taiwan. Blood samples were collected from each bear for hematological analysis and plasma biochemistry using a hematology analyzer. Parasites and pathogens were detected using a thin blood smear with Wright-Giemsa staining and polymerase chain reaction (PCR) assay. Additionally, macroscopic ectoparasites were collected from bears to detect blood parasites and other pathogens. Moreover, the relationships between the bear variables (sex, age, and occurrence of parasites or pathogens), ectoparasites, and infectious agents were also analyzed. RESULTS: In all, 21 wild bears (14 in YNP and 7 in DSY) were captured and released during the satellite tracking studies. Hematological analysis and plasma biochemistry indicated significant differences in white blood cells (WBC), segments, creatine kinase (CK), and lactate dehydrogenase (LDH) levels between foot snare and culvert-captured bears. Additionally, there were significant differences in total plasma protein (TPP), creatinine, Ca2+, Mg2+, and K+ levels between male and female bears. Moreover, pathogen-infected bears had significantly higher erythrocyte sedimentation rate (ESR; 30 min and 1 h) and globulin levels than uninfected bears. In total, 240 ticks were collected from 13 bears, among which eight adult tick species were identified, including Haemaphysalis flava, Haemaphysalis hystricis, Amblyomma testudinarium, Ixodes ovatus, Dermacentor taiwanensis, Haemaphysalis longicornis, Ixodes acutitarsus, Amblyomma javanense, and nymphs belonging to Haemaphysalis spp. PCR revealed that 13 (61.90%) and 8 (38.10%) bears harbored Hepatozoon ursi and Babesia DNA, respectively. Among the ticks examined, 157 (65.41%) and 128 (53.33%) samples were positive for H. ursi and Babesia, respectively. CONCLUSIONS: To the best of our knowledge, this is the first study to establish a database of the hematological and plasma profiles of wild Formosan black bears and investigate ectoparasite infestation and Hepatozoon and Babesia spp. INFECTION: In conclusion, these findings may serve as a reference for monitoring the health and population of locally endangered bears.

Analysis of doramectin in the serum of repeatedly treated pastured cattle used to predict the probability of cattle fever ticks (Acari: Ixodidae) feeding to repletion.

Analysis of doramectin concentration in blood serum of pastured cattle injected repeatedly (12 treatments) at two different dosage rates and 28-day intervals throughout the year was used to predict the probability that cattle fever ticks could successfully feed to repletion during the interval between any two consecutive treatments. Treatment at ~270 µg/kg indicated that serum doramectin concentration dropped below the baseline concentration estimated for tick survival (8 ppb) in 7 of the 12 treatments. However, the longest period between any two treatments during which the doramectin concentration remained below the 8 ppb baseline level for successful tick feeding was 15 days, making it virtually impossible for any ticks to reach ovipositional status prior to a subsequent treatment. At a dosage rate of ~540 µg/kg, the concentration dropped below the baseline tick survival level (8 ppb) only once, following the initial treatment, and the duration during which the concentration remained below the baseline level prior to the subsequent treatment was only 6 days. Thus, at the high dosage rate results indicated, with absolute certainty, that no ticks could successfully feed to repletion between any two consecutive treatments. Based on the data obtained in the study it was concluded that analysis of doramectin concentration in serum of treated animals would be a reliable predictor for assessing the probability that ticks could successfully develop to repletion. More importantly, results demonstrated that the trial policy, instituted by the Cattle Fever Tick Eradication Program, of repeatedly treating cattle with doramectin injections at 25-28 day intervals for eliminating cattle fever ticks would produce little or no risk of any viable ticks developing to repletion and re-infesting the field between treatment applications.

Effect of various levels of dietary Jatropha curcas seed meal on rabbits infested by the adult ticks of Hyalomma marginatum marginatum I. Animal performance, anti-tick feeding and haemogram.

The goal of this study was to investigate the use of Jatropha curcas seed meal (JCSM) in different levels as acaricide in diet of rabbits experimentally infested by Hyalomma marginatum marginatum then determining animal performance, anti-tick feeding and its effects on haemogram of rabbits. Thirty healthy mixed-breed rabbits were randomly divided into five equal groups. The first group was kept as a control fed soya bean meal (20%) as a source of protein. Groups from the second to the fifth fed diets contained 2.5%, 5%, 7.5% and 10% of JCSM instead of soya bean meal as a source of protein, respectively. Feeding and watering were given freely throughout the study. Animal performance for treatment groups were recorded from the 1st week up to the 6th week. Then each group divided into two subgroups, and the ticks were introduced to all of one subgroup and the other kept as control, following them until dropped at the end of the 8th week for all groups of the experiment. Feeding and reproductive performance of the adult tick females were determined. Blood samples were collected and analysed for haematological examination at the 0, 6th and 8th weeks post-treatment from all animals. Result revealed that rabbits received diets containing 5%, 7.5% and 10% had significantly (P < 0.05) lower growth performance than control and 2.5% JCSM groups. Feed intake and body weight gain decreased with increasing the level of JCSM in the diet. The result of anti-tick feeding observed that the highest percentage (60-90%) of rejection was recorded in 10% of JCSM group then the other treated groups (20-30%). Egg mass and reproductive index per female were marked increase (P < 0.001) in groups 2.5%, 5% and 7.5% of JCSM. Macrocytic normochromic anaemia was development after 8 weeks of treatment, which changed to microcytic normochromic anaemia after challenge of ticks in groups received 5%, 7.5% and 10% JCSM. Leukopenia, neutopenia and lymphopenia were noticed (P < 0.05) in all treated groups which more drop in the group received 7.5% JCSM. Also, monocytosis was recorded in 7.5% and 10% JCSM groups. In conclusion, JCSM could be use in the treatment of ectoparasites at level less than 10% in diet. Further investigations should be done to detoxification the Jatropha seed meal to decrease the level of its toxicity.

Allelic Variation in Protein Tyrosine Phosphatase Receptor Type-C in Cattle Influences Erythrocyte, Leukocyte and Humoral Responses to Infestation With the Cattle Tick Rhipicephalus australis.

The protein tyrosine phosphatase receptor type-C (PTPRC) gene encodes the common leukocyte antigen (CD45) receptor. CD45 affects cell adhesion, migration, cytokine signalling, cell development, and activation state. Four families of the gene have been identified in cattle: a taurine group (Family 1), two indicine groups (Families 2 and 4) and an African "taurindicine" group (Family 3). Host resistance in cattle to infestation with ticks is moderately heritable and primarily manifests as prevention of attachment and feeding by larvae. This study was conducted to describe the effects of PTPRC genotype on immune-response phenotypes in cattle that display a variable immune responsiveness to ticks. Thirty tick-naïve Santa-Gertrudis cattle (a stabilized composite of 5/8 taurine and 3/8 indicine) were artificially infested with ticks weekly for 13 weeks and ranked according to their tick counts. Blood samples were taken from control and tick-challenged cattle immediately before, then at 21 d after infestation and each subsequent week for 9 weeks. Assays included erythrocyte profiles, white blood cell counts, the percentage of cellular subsets comprising the peripheral blood mononuclear cell (PBMC) population, and the ability of PBMC to recognize and proliferate in response to stimulation with tick antigens in vitro. The cattle were PTPRC genotyped using a RFLP assay that differentiated Family 1 and 3 together (220 bp), from Family 2 (462 bp), and from Family 4 (486 bp). The PTPRC allele frequencies were Family 1/3 = 0.34; Family 2 = 0.47; Family 4 = 0.19. There was no significant association between PTPRC genotype and tick count. Each copy of the Family 1/3 allele significantly decreased total leucocyte count (WCC) and CD8+ cells. Increasing dosage of Family 2 alleles significantly increased red blood cell count (RCC), haematocrit (PCV), and haemoglobin (Hb) concentration in blood. Increasing dosage of the Family 4 allele was associated with increased WCC, reduced RCC, reduced PCV and reduced Hb. Homozygote Family 1/3 animals had consistently lower IgG1 in response to tick Ag than homozygote Family 2 animals. The PTPRC genotype influences the bovine immune response to ticks but was not associated with the observed variation in resistance to tick infestation in this study.

Evaluation on two types of paramyosin vaccines for the control of Haemaphysalis longicornis infestations in rabbits.

BACKGROUND: Haemaphysalis longicornis is an obligate hematophagous ectoparasite that transmits a variety of pathogens causing life-threatening diseases in humans and animals. Paramyosin (Pmy) is not only an invertebrate-specific myofibrillar protein but also an important immunomodulatory protein. Therefore, it is one of the ideal candidate antigens for vaccines. METHODS: We conducted two vaccine trials to evaluate the protective efficacy of Pmy recombinant protein (rPmy) and peptide vaccine (KLH-LEE). Each rabbit was immunized with three doses of rPmy or KLH-LEE adjuvanted with Freund's complete/incomplete at 500 µg/dose at 2-week intervals before challenge with 40 female H. longicornis/rabbit. PBS plus adjuvant, Trx or KLH was used as control group. The antibodies of rabbits were detected by ELISA. Then, female ticks were fed on the rabbits until detachment. RESULTS: ELISA results showed that both vaccines induced rabbits to produce antibodies. Compared with the Trx group, the engorgement weight, oviposition and hatchability of the rPmy group decreased by 8.87%, 26.83% and 38.86%, respectively. On the other hand, engorgement weight, oviposition and hatchability of female ticks in the KLH-LEE group correspondingly resulted in 27.03%, 53.15% and 38.40% reduction compared with that of the KLH group. Considering the cumulative effect of vaccination on the evaluated parameters, results showed 60.37% efficacy of the rPmy vaccine formulation and 70.86% efficacy in the KLH-LEE group. CONCLUSIONS: Pmy and particularly epitope LEE have potential for further development of an effective candidate vaccine to protect the host against tick infection. GRAPHIC ABSTARCT.

Rhipicephalus (Boophilus) microplus: clotting time in tick-infested skin varies according to local inflammation and gene expression patterns in tick salivary glands.

Ticks deposit saliva at the site of their attachment to a host in order to inhibit haemostasis, inflammation and innate and adaptive immune responses. The anti-haemostatic properties of tick saliva have been described by many studies, but few show that tick infestations or its anti-haemostatic components exert systemic effects in vivo. In the present study, we extended these observations and show that, compared with normal skin, bovine hosts that are genetically susceptible to tick infestations present an increase in the clotting time of blood collected from the immediate vicinity of haemorrhagic feeding pools in skin infested with different developmental stages of Rhipicepahlus microplus; conversely, we determined that clotting time of tick-infested skin from genetically resistant bovines was shorter than that of normal skin. Coagulation and inflammation have many components in common and we determined that in resistant bovines, eosinophils and basophils, which are known to contain tissue factor, are recruited in greater numbers to the inflammatory site of tick bites than in susceptible hosts. Finally, we correlated the observed differences in clotting times with the expression profiles of transcripts for putative anti-haemostatic proteins in different developmental stages of R. microplus fed on genetically susceptible and resistant hosts: we determined that transcripts coding for proteins similar to these molecules are overrepresented in salivary glands from nymphs and males fed on susceptible bovines. Our data indicate that ticks are able to modulate their host's local haemostatic reactions. In the resistant phenotype, larger amounts of inflammatory cells are recruited and expression of anti-coagulant molecules is decreased tick salivary glands, features that can hamper the tick's blood meal.

Systemic alterations of bovine hemostasis due to Rhipicephalus (Boophilus) microplus infestation.

The tick Rhipicephalus (Boophilus) microplus is a hematophagous ectoparasite that causes considerable economic losses to cattle breeding. Although R. microplus saliva contains several molecules that interfere with the blood coagulation process, so far the systemic alterations in the host hemostatic system have not been described. This study aims to determine if R. microplus infestation induces any disturbance to the host's hemostatic system. To address these questions, six calves were experimentally infested with 20,000 R. microplus larvae and their blood was collected before and 7, 14 and 21 days post-infestation. Collagen and ADP-induced platelet aggregation as well as coagulation (activated partial thromboplastin time and prothrombin time) decreased in infested bovines. Platelet blood count and fibrinogen increased during the course of infestation, probably as a compensatory response. These alterations may play a role in host health status, and show that the host cannot fully counteract the tick anti-hemostatic mechanisms.

[Two cases of Crimean-Congo haemorrhagic fever (CCHF) in two tourists in Senegal in 2004]. / Deux cas de fièvre hémorragique de Crimée-Congo (FHCC) contractée au Sénégal, en 2004, par des résidentes temporaires.

Two cases of Crimean-Congo haemorrhagic fever (CCHF) occurred in two French tourists during their visit in Senegal in November 2004. Febrile and hemorrhagic syndrome with ulorrhagia, petechiae, haematemesis, haematomas associated with biological signs of disseminated intramuscular coagulation were observed. For the first case who had a medical evacuation to France before diagnosis, Crimean-Congo virus infection was revealed by laboratory tests performed by the National Reference Center for Hemorrhagic Fevers (NRCHF, Institut Pasteur, Lyon) and secondly by the Centre de Référence OMS sur la Recherche des Arbovirus et des virus des Fièvres Hémorragiques (CRORA) in the Dakar Pasteur Institute (DPI). The second case diagnosed by the CRORA died after clinical deterioration with liver failure and severe haemorrhages. Healthcare workers and family members who had contact with tissue or blood from patients were followed up after the putative exposure either in France or in Senegal.

Comparative IgG recognition of tick extracts by sera of experimentally infested bovines.

Enzyme-linked immunosorbent assay (ELISA) and Western blot were used to investigate the pattern of antibody responses of six bovines infested twelve times with Rhipicephalus (Boophilus) microplus (Canestrini, 1887) (Acari: Ixodidae) (six heavy infestations followed by six light infestations) against salivary gland, gut and larvae extracts. During heavy infestations, bovine IgG levels were shown to be higher, and a decrease in the number and weight of ticks that completed the parasitic cycle was observed. The pattern changed starting from the seventh infestation, showing a decrease in IgG levels. An initial increase followed by a significant decrease in the proportion of ticks that completed the parasitic cycle was also observed from the seventh infestation. The number of molecules recognized by Western blot was higher from sera collected following heavy infestations than after light infestations, although a great variation in the profiles detected could be seen when the bovines were compared. These results indicate that IgG responses to different tick antigens may not be generally associated with bovine resistance, and that infestation levels modulate the magnitude of humoral responses and possibly the immune mechanisms in the natural acquisition of tick resistance.

Molecular Detection of Theileria annulata in Cattle from Different Regions of Punjab, Pakistan, by Using Recombinase Polymerase Amplification and Polymerase Chain Reaction.

Piroplasmosis is one of the most important diseases of livestock, constraining optimal production and leading to economic loss. This study was carried out to detect Theileria annulata by using 2 different molecular techniques: recombinase polymerase amplification (RPA) and conventional polymerase chain reaction (PCR). Blood samples were collected from 274 ticks infesting asymptomatic cattle from several counties in the Chakwal, Faisalabad, and Jhang districts of Punjab Province in Pakistan by using FTA cards. After extraction of genomic DNA, each sample was subjected to RPA optimized to amplify a 281-bp fragment of the Enolase gene for T. annulata. The specificity of the test was confirmed using positive DNA samples of related piroplasm species, whereas the analytical sensitivity was calculated using different serial dilutions of a long fragment of the same gene. The RPA results were positive for 48 (17.51%) of 274 samples. All 274 samples were screened using conventional PCR, and 21 (7.66%) samples were positive for T. annulata. All the samples that were RPA positive but PCR negative were sequenced, which confirmed the results of RPA. The highest positive rate was found in Chakwal district, followed by Faisalabad and Jhang districts. This study demonstrates the application of highly sensitive and specific rapid diagnostic methods for T. annulata to a regional screening program. This is the first report of tick-borne disease from Pakistan by using RPA.

Patterns of Midichloria infection in avian-borne African ticks and their trans-Saharan migratory hosts.

BACKGROUND: Ticks are obligate haematophagous ectoparasites of vertebrates and frequently parasitize avian species that can carry them across continents during their long-distance migrations. Ticks may have detrimental effects on the health state of their avian hosts, which can be either directly caused by blood-draining or mediated by microbial pathogens transmitted during the blood meal. Indeed, ticks host complex microbial communities, including bacterial pathogens and symbionts. Midichloria bacteria (Rickettsiales) are widespread tick endosymbionts that can be transmitted to vertebrate hosts during the tick bite, inducing an antibody response. Their actual role as infectious/pathogenic agents is, however, unclear. METHODS: We screened for Midichloria DNA African ticks and blood samples collected from trans-Saharan migratory songbirds at their arrival in Europe during spring migration. RESULTS: Tick infestation rate was 5.7%, with most ticks belonging to the Hyalomma marginatum species complex. Over 90% of Hyalomma ticks harboured DNA of Midichloria bacteria belonging to the monophylum associated with ticks. Midichloria DNA was detected in 43% of blood samples of avian hosts. Tick-infested adult birds were significantly more likely to test positive to the presence of Midichloria DNA than non-infested adults and second-year individuals, suggesting a long-term persistence of these bacteria within avian hosts. Tick parasitism was associated with a significantly delayed timing of spring migration of avian hosts but had no significant effects on body condition, whereas blood Midichloria DNA presence negatively affected fat deposits of tick-infested avian hosts. CONCLUSIONS: Our results show that ticks effectively transfer Midichloria bacteria to avian hosts, supporting the hypothesis that they are infectious to vertebrates. Bird infection likely enhances the horizontal spread of these bacteria across haematophagous ectoparasite populations. Moreover, we showed that Midichloria and tick parasitism have detrimental non-independent effects on avian host health during migration, highlighting the complexity of interactions involving ticks, their vertebrate hosts, and tick-borne bacteria.

Metabolic differentiation of early Lyme disease from southern tick-associated rash illness (STARI).

Lyme disease, the most commonly reported vector-borne disease in the United States, results from infection with Borrelia burgdorferi. Early clinical diagnosis of this disease is largely based on the presence of an erythematous skin lesion for individuals in high-risk regions. This, however, can be confused with other illnesses including southern tick-associated rash illness (STARI), an illness that lacks a defined etiological agent or laboratory diagnostic test, and is coprevalent with Lyme disease in portions of the eastern United States. By applying an unbiased metabolomics approach with sera retrospectively obtained from well-characterized patients, we defined biochemical and diagnostic differences between early Lyme disease and STARI. Specifically, a metabolic biosignature consisting of 261 molecular features (MFs) revealed that altered N-acyl ethanolamine and primary fatty acid amide metabolism discriminated early Lyme disease from STARI. Development of classification models with the 261-MF biosignature and testing against validation samples differentiated early Lyme disease from STARI with an accuracy of 85 to 98%. These findings revealed metabolic dissimilarity between early Lyme disease and STARI, and provide a powerful and new approach to inform patient management by objectively distinguishing early Lyme disease from an illness with nearly identical symptoms.

Dog tick-borne diseases in Sicily.

In Sicily many tick borne diseases are endemic, in particular way those that see like main carrier ticks that prefer, for their vital cycle, climatic conditions characterized by high temperatures and a warmth-humid atmosphere. The more important pathologies transmitted by ticks causing diseases in dogs are babesiosis and ehrlichiosis. Borrelia burgdorferi, Anaplasma phagocytophilum, Rickettsia conorii, Coxiella burnetii and tick transmitted encephalitis virus assume particular relevance because they are agents of zoonosis. Our centre, C.R.A.Ba.R.T, have conducted many researches and carried out many tests for diagnostic aim in order to estimate the spread of the main tick borne diseases in Sicilians' dogs. A study lead on 342 dogs has evidenced seroprevalence for Babesia canis, Ehrlichia canis and Rickettsia respective of 5.17%, 21.70% and 53.43%. A study on zoonotic agent seroprevalences in dogs gave the following percentages: C. burnetii 31.50%, R. conorii 73.60% and A. phagocytophilum 32.80%. The data carried out from IZS Sicily diagnostic service on 5,634 tests done in 2004-2005, confirm the experimental results on the presence of B. canis, E. canis, R. conorii, A. phagocytophilum, C. burnetii, Rickettsia spp., Anaplasma spp. and Ehrlichia spp. in all the Sicilian areas.

NT-proBNP and cardiac troponin I concentrations in dogs with tick paralysis caused by Ixodes holocyclus.

OBJECTIVE: The purpose of this study was to determine through measurement of cardiac biomarkers whether there was cardiac involvement in dogs infested with Ixodes holocyclus. METHODS: Dogs with tick paralysis and no-mild (group 1; n = 44) or moderate-severe respiratory compromise (group 2; n = 36) and a control group of dogs (n = 31) were enrolled. Plasma N-terminal pro-B-type natriuretic peptide (NT-proBNP), serum cardiac troponin I (cTnI) and serum creatinine concentrations were determined. For most of the affected dogs SpO2 was determined. RESULTS: SpO2 readings did not differ between groups 1 and 2. Three animals in group 2 had an SpO2 reading <90%. NT-proBNP concentrations were lower in both groups 1 and 2 compared with the control group. There was no difference in cTnI concentrations among groups, although they were elevated in four dogs, including the three dogs in group 2 with SpO2 readings <90%. Creatinine concentrations were within the reference interval for all dogs, but did differ among the groups, with control dogs having the highest values, followed by group 1 and then group 2. CONCLUSION: This study did not detect significant cardiac involvement in dogs with tick paralysis induced by I. holocyclus. Evidence for reduced preload in dogs with tick paralysis was provided by lower NT-proBNP concentrations compared with control dogs. Severe hypoxaemia may not be a significant component of the clinical picture in many of the dogs presenting with tick paralysis. Dogs with severe hypoxaemia may have loss of cardiomyocyte integrity, reflected by elevated cTnI concentrations.

Persistent Ehrlichia ewingii infection in dogs after natural tick infestation.

BACKGROUND: Ehrlichia ewingii, which causes disease in dogs and people, is the most common Ehrlichia spp. infecting dogs in the United States, but little is known about how long E. ewingii infection persists in dogs. HYPOTHESIS/OBJECTIVES: To evaluate the persistence of natural infection with E. ewingii in dogs. ANIMALS: Four Class A Beagles; no previous exposure to ticks or tick-borne infectious agents. METHODS: Dogs were exposed to ticks by weekly walks through tick habitat in north central Oklahoma; dogs positive for infection with Ehrlichia spp. by sequence-confirmed PCR and peptide-specific serology were evaluated for 733 days (D). Whole blood was collected once weekly for PCR, and serum was collected once monthly for detection of antibodies to Ehrlichia canis (peptide p16), Ehrlichia chaffeensis (indirect fluorescence antibody [IFA] and variable-length PCR target [VLPT]), and E. ewingii (peptide p28). RESULTS: All dogs (4/4) became infected with Ehrlichia spp. as evidenced by seroconversion on IFA to E. chaffeensis (4/4); PCR detection of E. ewingii (4/4) and E. chaffeensis (2/4) DNA using both nested and real-time assays; and presence of specific antibodies to E. ewingii (4/4) and E. chaffeensis (2/4). Infection with E. chaffeensis was not detected after D55. Intermittent E. ewingii rickettsemia persisted in 3 of 4 dogs for as long as 733 days. CONCLUSIONS AND CLINICAL IMPORTANCE: Our data demonstrate that dogs infected with E. ewingii from tick feeding are capable of maintaining infection with this pathogen long-term, and may serve as a reservoir host for the maintenance of E. ewingii in nature.

Amblyomma maculatum Feeding Augments Rickettsia parkeri Infection in a Rhesus Macaque Model: A Pilot Study.

Rickettsia parkeri is an emerging eschar-causing human pathogen in the spotted fever group of Rickettsia and is transmitted by the Gulf coast tick, Amblyomma maculatum. Tick saliva has been shown to alter both the cellular and humoral components of the innate and adaptive immune systems. However, the effect of this immunomodulation on Rickettsia transmission and pathology in an immunocompetent vertebrate host has not been fully examined. We hypothesize that, by modifying the host immune response, tick feeding enhances infection and pathology of pathogenic spotted fever group Rickettsia sp. In order to assess this interaction in vivo, a pilot study was conducted using five rhesus macaques that were divided into three groups. One group was intradermally inoculated with low passage R. parkeri (Portsmouth strain) alone (n = 2) and another group was inoculated during infestation by adult, R. parkeri-free A. maculatum (n = 2). The final macaque was infested with ticks alone (tick feeding control group). Blood, lymph node and skin biopsies were collected at several time points post-inoculation/infestation to assess pathology and quantify rickettsial DNA. As opposed to the tick-only animal, all Rickettsia-inoculated macaques developed inflammatory leukograms, elevated C-reactive protein concentrations, and elevated TH1 (interferon-γ, interleukin-15) and acute phase inflammatory cytokines (interleukin-6) post-inoculation, with greater neutrophilia and interleukin-6 concentrations in the tick plus R. parkeri group. While eschars formed at all R. parkeri inoculation sites, larger and slower healing eschars were observed in the tick feeding plus R. parkeri group. Furthermore, dissemination of R. parkeri to draining lymph nodes early in infection and increased persistence at the inoculation site were observed in the tick plus R. parkeri group. This study indicates that rhesus macaques can be used to model R. parkeri rickettsiosis, and suggests that immunomodulatory factors introduced during tick feeding may enhance the pathogenicity of spotted fever group Rickettsia.

Blood eosinophil and basophil responses in guinea pigs parasitized by Amblyomma americanum ticks.

Guinea pigs sensitized to tick feeding by a previous infestation express immune resistance when challenged. This state is associated with large accumulations of eosinophils, and especially of basophils, at cutaneous sites of tick feeding. The present study shows that primary and secondary tick feeding stimulate blood eosinophilia and basophilia that reflect the infiltration of these cells in the skin. Primary feeding of Amblyomma americanum larvae resulted in a mean tick yield of 81%, mean tick weight of 0.91 mg and a feeding period of 3-5 days. Blood eosinophils and basophils began to rise on day 2 and reached peak 3-fold increases on day 4, the time of maximum tick drop-off. Secondary feedings 26 days later resulted in a mean tick yield of 45% (52% tick rejection), mean tick weight of 0.71 mg (22% weight decrease) and a mean feeding period of 4-6 days with maximum detachment on day 5. Prior to secondary tick feeding, both eosinophil and basophil levels were greater than in tick-free animals. Eosinophil levels peaked at day 4 to a level three times greater than in primary tick hosts. Basophil levels in secondary hosts exhibited minor elevations until day 6 when they were increased significantly, reaching a peak level of 33% higher than that observed in primary feedings. This study demonstrates that blood eosinophils and basophils respond quickly and significantly to primary tick feeding and undergo secondary responses of greater magnitude in response to subsequent tick challenge. However, challenge feeding resulted in a large blood eosinophilia with a modest blood basophilia. This may reflect the balance between bone marrow production and cutaneous accumulation of these cells at sites of tick rejection. Recruitment of eosinophils and basophils from the blood to the tissue site of tick feeding may contribute to the tick resistance mechanism.

Isotopes as host blood markers to measure blood intake by feeding ticks (Acari: Ixodidae).

We have modified an experimental technique in which two different isotopes (Cr51 and I125) were used simultaneously as blood markers for determining the amount of blood removed by feeding ticks. This method enables us to measure separately and directly the volumes of red blood cells and plasma in ticks and to calculate the ratio between the two parameters (concentrating ability). The concentration of red blood cells in nymphs of Amblyomma hebraeum Kock and Rhipicephalus appendiculatus Neumann and in females of these species plus in those of Hyalomma truncatum Kock were found to be twice the amount measured in the labeled blood of goats. Concentrations of plasma in the ticks were approximately two-thirds of that measured in samples taken directly from these same goats. Small nonsignificant differences between the ability of various tick species to concentrate red blood cells and dilute host plasma were found. The advantage of this method is compared with that of the colorimetric and gravimetric methods, and its potential use is discussed.