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PubMed1997

Vaccines against hemoparasitic diseases in Israel with special reference to quality assurance.

Four vaccines against hemoparasitic diseases (anaplasmosis, babesiosis and theileriosis) and a vaccine against besnoitiosis are currently used in Israel. These vaccines contain live attenuated parasites derived from cell culture (Theileria annulata and Besnoitia besnoiti) or from blood of infected, splenectomized calves (Babesia bigemina, B. bovis and Anaplasma centrale). Cryopreserved master seed is used to initiate production of the vaccines. Quality control performed during the preproduction period is particularly important with blood-derived vaccines. Post production quality control comprises sterility, potency (viability of immunizing organisms), safety (degree of attenuation) and efficacy (capacity to protect against virulent parasite stock). All vaccines are stored and dispatched to the field in a concentrated frozen state. The culture-derived vaccines are safe for all varieties of cattle, regardless of age or physiological condition, whereas the blood-derived vaccines are recommended mainly for young cattle, the age limit varying with the type of vaccine and breed of cattle. The viability of T. annulata infected cells in the anti-theilerial vaccine is tested after thawing by in vitro plating efficiency and the infectivity of blood-derived vaccines is tested by titration in susceptible cattle.

Anaplasma↗
PubMed1983

Acquired cellular responsiveness in cattle cleared of Anaplasma marginale 28 months earlier.

The leukocyte migration inhibition test (LMIT) was used to assess cell-mediated immune responses of cattle against Anaplasma marginale. Leukocytes from 7 of 11 cows that had been cleared of the carrier state by antibiotic therapy 28 months earlier were inhibited from their normal migration by A. marginale antigens (P less than 0.001). After reexposure to virulent organisms, all animals were positive to the LMIT. There was not a significant relationship (r = 0.49) between LMIT values before and after reexposure. Results suggest that animals free of anaplasmosis for more than 2 years are capable of cell-mediated immune responses, despite decreased in vitro responsiveness at the time of reexposure. Acquired cellular responses did not prevent reinfection as all animals became A. marginale carriers.

Anaplasma↗
PubMed1982

A case of apparent suppression of Anaplasma marginale infection by eperythrozoonosis (Eperythrozoon teganodes).

The apparent suppression of Anaplasma marginale infection by Eperythrozoon teganodes in a splenectomized calf has been reported. A splenectomized calf, inoculated with 500 ml of blood having 23% erythrocytes infected with A. marginale, developed eperythrozoonosis on the fourth day post inoculation. A. marginale parasitaemia remained very low during the patent eperythrozoonosis. A. marginale parasites started to increase in number only after E. teganodes infection had been controlled with neoarsphenamine. A splenectomized calf treated identically, but not showing E. teganodes parasites in the peripheral blood, developed clinical anaplasmosis and fulminant parasitaemia within 3-4 days post inoculation.

Anaplasmataceae Infections↗
PubMed1988

A sensitive ELISA technique for the diagnosis of Anaplasma marginale infections.

A sensitive enzyme-linked immunosorbent assay (ELISA) technique using a horse radish peroxidase conjugate is described for measuring Anaplasma marginale antibodies in bovine serum. This technique utilizes two antigen preparations, a 'negative' antigen derived from an animal prior to infection and a 'positive' antigen derived from A. marginale-infected red cells from the same animal following infection. This markedly reduces cross-reactions which are a result of isoantigens. Absorbance values obtained using the 'negative' antigen are subtracted from those obtained using the 'positive' antigen to give a net figure. Of 100 A. marginale-positive sera tested no false negative results were obtained. All 11 animals maintained tick-free after initial diagnosis of naturally transmitted anaplasmosis were positive 3 years later, 15 A. marginale-infected animals maintained with ticks were positive 27 months after initial infection and a further 26 animals infected with A. marginale by blood inoculation were positive 3 months later. Three per cent of negative sera, 2% of B. bovis and 4% of B. bigemina-positive sera gave positive reactions.

Anaplasma↗
PubMed1989

Infectivity of cryopreserved Babesia bovis, Babesia bigemina and Anaplasma centrale for cattle after thawing, dilution and incubation at 30 degrees C.

Blood containing either Babesia bovis, Babesia bigemina or Anaplasma centrale was mixed with an equal volume of 3 M glycerol in phosphate-buffered saline with or without glucose and then stored in liquid nitrogen for 2-30 days. After being thawed, the parasitized blood was subjected to various procedures, including dilution up to 1000-fold followed by incubation at 30 or 4 degrees C for 8 h, before infectivity of the parasites was tested in a total of 70 cattle. The results showed that the blood cryopreserved with glycerol remained highly infective after thawing, despite dilution and incubation for 8 h at 30 degrees C. The results have practical application in the use of frozen, live vaccines against bovine babesiosis and anaplasmosis.

Anaplasma↗
PubMed1994

Chemotherapy and delivery systems: haemoparasites.

Chemotherapy of haemoparasitic diseases in domestic animals is dependent on a limited number of compounds, many of which are chemically closely related. In this review, a summary is given of each of the drugs currently available for treatment and prophylaxis of trypanosomosis and the tick-borne diseases theileriosis, babesiosis, anaplasmosis and cowdriosis. In contrast to the situation with the drugs used for tick-borne diseases, drug resistance appears to be becoming an increasing problem associated with the compounds used for trypanosomosis. The literature that has been reviewed, therefore, is that which relates to the methods used to identify and quantify drug resistance in trypanosome populations, reports of resistance to trypanocides, and cross-resistance between trypanosome populations, reports of resistance to trypanocides, and cross-resistance between trypanocides. The possible reason(s) for the apparent lack of development of resistance to the compounds used for treatment of tick-borne diseases is also discussed. Local toxicity at the site of injection is a problem that is particularly associated with many of the trypanocides when used on a long-term basis in individual animals. Various alternative preparations of the currently used trypanocides therefore have been evaluated in an attempt to reduce this toxicity, and are summarised. Finally, future developments in haemoparasitic chemotherapy are considered and, for trypanosomosis, highlight the importance of integrating chemotherapeutic and chemoprophylactic programmes with control of the vector when drug resistance becomes a significant constraint.

Anaplasmosis↗
PubMed1984

The effect of Anaplasma marginale on the glycolytic pathway in bovine erythrocytes.

The glycolytic process in non-infected and A. marginale-infected bovine erythrocytes was studied. The study included four enzymes: hexokinase, phosphofructokinase, pyruvic kinase and lactic dehydrogenase, and the intracellular concentrations of glucose, ATP and 2,3 diphosphoglyceric acid (DPG). The activities of phosphofructokinase and lactic dehydrogenase found in non-infected bovine erythrocytes were fifty times greater than the previously reported values. Glucose and DPG concentrations, hexokinase, pyruvic kinase and lactic dehydrogenase specific activities did not change significantly during experimental anaplasmosis. Phosphofructokinase activity in A. marginale-infected erythrocytes increased to values 300% of those found in intact bovine red cells. ATP concentration in infected erythrocytes decreased to values of 40% of those found in non-infected cells: from 1.60 +/- 0.40 to 0.70 +/- 0.08 mumole per gram of hemoglobin.

2,3-Diphosphoglycerate↗
PubMed1990

Comparison of a DNA probe, complement-fixation and indirect immunofluorescence tests for diagnosing Anaplasma marginale in suspected carrier cattle.

Most estimates of the prevalence of anaplasmosis have been based on serologic data using the complement-fixation (CF) and/or card agglutination tests. Since these tests are considered to be only about 50 percent reliable for detecting carrier cattle in enzootically stable herds, the need for more sensitive diagnostic tests is widely recognized. The objective in the present study was to compare the sensitivity of the CF test with that of the indirect immunofluorescence (IIF) test and a recently developed DNA probe in determining the prevalence of Anaplasma marginale infection in cattle from an enzootic area. The study herd consisted of 52 8-month-old steers and 13 3-year-old cows of mixed beef breed. All cattle were initially tested for this comparative purpose. All but one animal (one that was a positive reactor as assessed by all three tests, and served as a positive control), were treated with long-acting oxytetracycline in an attempt to clear any carrier infections. Each animal was then retested at 1 month and 2 months post-treatment (PT), in an effort to determine if the DNA probe could be used to evaluate the effectiveness of the drug. Six of the 65 (9.2%) initial serum samples were CF positive. In contrast, 60 (92.3%) and 64 (98.5%) of the samples were positive as assessed with the IIF test and the DNA probe, respectively. The DNA hybridization reactions varied in intensity within the sample population indicating different individual levels of infection. The DNA probe hybridized with two samples taken at 1 month PT, and with two different samples taken at 2 months PT. The mean IIF titers were reduced at both the 1 month and 2 month sampling times. These results suggest that the drug did not eliminate infections in all cattle. Some may have been cleared, but, in any event, the drug did reduce the level of infections below the sensitivity of the DNA probe and interrupted continuity of stimulation of antibody. Therefore, the DNA probe and the IIF test appear to be considerably more sensitive in detecting carrier infections than the CF test, and should be considered in future epidemiologic studies.

Anaplasma↗
PubMed1990

Development of a recombinant Anaplasma marginale DNA probe.

An Anaplasma marginale DNA probe has been developed by using an improved method for the isolation of genomic DNA. Purified genomic A. marginale DNA from the St. Croix isolate was partially digested with Sau 3A1 into fragments (greater than or equal to 5.0 kb). The restriction fragments were cloned using standard techniques in the pBR322 vector and used to transform E. coli (DH5) host cells. The recombinant A. marginale DNA library was screened by the colony lifting procedure. Colonies containing plasmids with A. marginale DNA inserts were identified by hybridization with a genomic A. marginale DNA radiolabeled probe (32P). Seven recombinant A. marginale DNA probes were evaluated by dot-blot in vitro hybridization assays to identify candidates as diagnostic tools in bovine anaplasmosis studies. Specificity and sensitivity experiments were carried out by using heterologous and homologous DNAs. The heterologous panel contained bovine DNA (WBC) and blood parasites DNA from Babesia bovis (Bb), Babesia bigemina (Bbi), Eperythrozoon suis (Es) and Eperythrozoon wenyoni (Ew). The homologous DNA panel included A. marginale DNAs of 12 different isolates which were isolated in the Caribbean, Mexico, and the U.S.A. The selected diagnostic probe was identified as pSt. Croix A1, and labeled with 32P by using in vitro nick translation and random primer techniques. The pSt. Croix A1 probe demonstrated 100% specificity and high sensitivity by hybridization in dot blotting and Southern blotting. The probe can detect 500-1000 infected erythrocytes per microliters which corresponds to a parasitemia of less than 0.01%. The A. marginale DNA insert was approximately 6.4 kb in size and a partial restriction map has been constructed.

Anaplasma↗
PubMed1993

Antigen profiles of Anaplasma ovis and A. mesaeterum and cross infection trials with them and A. marginale.

Antigen profiles of Anaplasma ovis and A. mesaeterum were analysed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting, and cross infection trials were performed by A. ovis challenge to A. mesaeterum- or A. marginale-inoculated goats and A. marginale challenge to an A. ovis-inoculated calf. Antigen analysis showed unique and common proteins of A. ovis and A. mesaeterum and identified a major 38 kDa protein to be a dominant immunogen bearing two epitopes common to Anaplasma species. An epitope specific to A. ovis and A. marginale, absent from A. mesaeterum, was recognised on the major protein of A. ovis. Pre-inoculation of goats with A. mesaeterum induced continuous antibody response with low parasitemia and partial protection against A. ovis challenge, as demonstrated by lower peak parasitemia and normal body temperature compared with values for goats inoculated with A. ovis alone. Pre-inoculation of goats with A. marginale and a calf with A. ovis induced weak or no antibody response without parasitemia, and was of no effect for preventing anaplasmosis by heterologous challenge. These results suggest that the development of parasitemia is essential for inducing continuous antibody by which cross protection would be possible.

Anaplasma↗
PubMed2006

Etiology of tick-borne febrile illnesses in adult residents of North-Eastern Poland: report from a prospective clinical study.

Febrile tick-borne diseases can be caused by different pathogens. The study objective was to prospectively determine the etiology of infection among adults exposed to tick bite and to detect acute human granulocytic anaplasmosis (HGA), a recently emerging infection, in north-eastern Poland. We evaluated 68 patients (34 males and 34 females, mean age 44.2+/-15 years) who had fever within 4 weeks after a tick bite. Paired sera were tested for antibodies against tick-borne encephalitis virus, Borrelia burgdorferi and Anaplasma phagocytophilum. Microscopic examination of EDTA whole blood and nested PCR targeting A. phagocytophilum 16S rDNA gene fragment were carried out at enrolment. Tick-borne infections were diagnosed in 57 individuals (84%). Sixty-four patients were hospitalised. Tick-borne encephalitis (TBE) was revealed in 49 cases; in 39 patients as a single infection, in three patients concurrent with erythema migrans or other Lyme borreliosis symptoms, in five patients concurrent with probable or possible Lyme borreliosis, and in two patients concurrent with A. phagocytophilum infection. Diagnosis of HGA was confirmed in one patient by PCR and in another one by seroconversion. In two further individuals, A. phagocytophilum infection was confirmed by immunofluorescence (antibody titers > or = 1:128), which, however, does not fulfil the ESCAR (European Society of Clinical Microbiology and Infectious Diseases Study Group on Coxiella, Anaplasma, Rickettsia, and Bartonella) criteria for HGA case definition. Lyme borreliosis was diagnosed in 16 patients; in six of them as a single infection. Both confirmed HGA cases and seropositive individuals had A. phagocytophilum infection concurrent either with Lyme borreliosis (two cases) or with TBE (two cases). The clinical course was severe in three and moderate to mild in the remaining TBE cases, as well as in all Lyme borreliosis and HGA cases. TBE was found the most prevalent disease among adults with febrile illnesses occurring after a tick bite in north-eastern Poland. Concurrent tick-borne infections were frequent with multiple pathogens involved. Two confirmed acute HGA cases (fulfilling the European case definition criteria defined by ESCAR) were detected for the first time in a prospective manner in Poland.

Adolescent↗
PubMed2006

Herbal usage and informant consensus in ethnoveterinary management of cattle diseases among the Kikuyus (Central Kenya).

For most smallholder farmers in Kenya conventional veterinary drugs have become very expensive and therefore unaffordable, causing them to seek low cost alternatives that are rarely documented in most ethnobiological studies. This study surveyed the utilisation of traditional herbal preparations in managing cattle ailments in Central Kenya with the aim of providing a comprehensive ethnobotanical profile and the most important plant species that may warrant scientific validation for efficacy and commercial utilisation. Using semi-structured questionnaires and detailed discussions with smallholder farmers, a total of 40 plant species in 26 families were found to be useful in traditional management of various cattle ailments in this region. Two plant families were particularly frequent in usage: Asteraceae and Lamiaceae, while the most utilised plant species were found to be Synadenium compactum N.E.Br. (Euphorbiaceae), Solanecio manii (Hook.f.) C. Jeffrey (Asteraceae) and Senna didymobotrya (Fresen.) Irwin and Barneby (Caesalpinaceae). Informant consensus was particularly high in managing anaplasmosis, East coast fever and ectoparasites. Such plant species become key target in efficacy tests and for development of commercial veterinary botanicals. The usage of some of the species is unfortunately unsustainable as some of the species are rare or endangered hence the need for conservation strategies to be undertaken.

Anaplasmosis↗
PubMed2004

Anaplasma infection in free-ranging Iberian red deer in the region of Castilla-La Mancha, Spain.

Organisms in the genus Anaplasma are obligate intracellular pathogens that multiply in both vertebrate and invertebrate hosts. The type species, Anaplasma marginale, causes bovine anaplasmosis and infects erythrocytes of the vertebrate host and undergoes a complex developmental cycle in ticks which serve as biological vectors. Infected cattle, wild ruminants and ticks can all serve as reservoirs of A. marginale. In this study, hunter killed Iberian red deer (Cervus elaphus hispanicus) from the region of Castilla-La Mancha in southwestern Spain were tested for Anaplasma infection. We found that 10% of the deer examined were seropositive for Anaplasma. Three A. marginale strains were subsequently obtained from salivary glands of Hyalomma marginatum that were removed from these deer, and the sequence of the major surface protein (msp)4 gene was determined for each strain and used for phylogenetic studies. Maximum parsimony analyses of msp4 sequences from H. marginatum ticks in comparison with New World cattle and bison isolates reported previously, suggested different origins for these Spanish A. marginale strains. The results of this study demonstrated that Iberian red deer are naturally infected with Anaplasma, and may therefore serve as a wildlife reservoir of the pathogen. Although the link between deer infection and the strains of A. marginale identified in ticks was not established, H. marginatum and Rhipicephalus bursa were identified as potential biological vectors for A. marginale in this region and may effect transmission of A. marginale between deer and cattle populations.

Amino Acid Sequence↗
PubMed2005

Anaplasma marginale: lack of cross-protection between strains that share MSP1a variable region and MSP4.

In Mexico, there are no commercial alternatives for the immunoprophylaxis of bovine Anaplasmosis, a disease responsible for great economic losses. Blood derived Anaplasma marginale used for immunizing susceptible cattle has shown promising results for homologous protection and controversial results against unrelated strains. The present study examined, under controlled conditions, the cross-protective potential of an immunogen composed of blood derived A. marginale of three strains against challenge with strains not included in the immunogens. Groups 1 and 2 were immunized with blood derived Anaplasma from strains Mexico, Morelos and Yucatan, group 4 with strains Morelos, Veracruz and Yucatan, two more groups (2 and 5) of equal conditions were inoculated with an adjuvant alone. Groups 1, 4 and 5 were challenged with Mexico strain; groups 2 and 3 were challenge-inoculated with strain Veracruz; groups 3 and 5 with strains Veracruz and Mexico as controls. Only animals in group 1, immunized and challenged with strain Mexico showed adequate protection. Both groups challenged with strains not included in the immunogens developed poor protection, while all the controls had to be treated to prevent death.

Anaplasma marginale↗
PubMed2006

Analysis of world strains of Anaplasma marginale using major surface protein 1a repeat sequences.

Anaplasma marginale is a tick-borne pathogen of cattle that causes the disease bovine anaplasmosis worldwide. Major surface proteins (MSPs) are involved in host-pathogen and tick-pathogen interactions and have been used as markers for the genetic characterization of A. marginale strains and phylogenetic studies. MSP1a is involved in the adhesion and transmission of A. marginale by ticks and varies among geographic strains in the number and sequence of amino-terminal tandem repeats. The aim of this study was to characterize the genetic diversity of A. marginale strains collected from countries in North and South America, Europe, Asia, Africa and Australia, inclusive of all continents. In this study, we characterized 131 strains of A. marginale using 79 MSP1a repeat sequences. These results corroborated the genetic heterogeneity of A. marginale strains in endemic regions worldwide. The phylogenetic analyses of MSP1a repeat sequences did not result in clusters according to the geographic origin of A. marginale strains but provided phylogeographic information. Seventy-eight percent of the MSP1a repeat sequences were present in strains from a single geographic region. Strong (> or =80%) support was found for clusters containing sequences from Italian, Spanish, Chinese, Argentinean and South American strains. The phylogenetic analyses of MSP1a repeat sequences suggested tick-pathogen co-evolution and provided evidence of multiple introductions of A. marginale strains from various geographic locations worldwide. These results contribute to the understanding of the genetic diversity and evolution of A. marginale and tick-pathogen interactions.

Amino Acid Sequence↗
PubMed2006

Detection of Ehrlichia chaffeensis in Brazilian marsh deer (Blastocerus dichotomus).

Ehrlichia chaffeensis was detected for the first time in blood samples from Brazilian marsh deers (Blastocerus dichotomus) captured in the marshes of Parana River in Southeast Brazil in 1998. Seven EDTA-blood samples from deers were analyzed by PCR and nested PCR for presence of Ehrlichia chaffeensis, Ehrlichia ewingii, Ehrlichia canis, Neoriickettsia risticii, Anaplasma phagocytophilum and Anaplasma marginale. Three samples showed positive reactions for E. chaffeensis and Anaplasma marginale. None contained detectable A. phagocytophilum, E. ewingii, E. canis or Neorickettsia risticii DNA. In Brazil, the wild marsh deer may be a natural reservoir of the agents that cause human monocytotropic ehrlichiosis and ruminant erythrocytic anaplasmosis.

Anaplasma marginale↗
PubMed2001

Identification of antigenic differences that discriminate between cattle vaccinated with Anaplasma centrale and cattle naturally infected with Anaplasma marginale.

Monoclonal antibodies were raised against the vaccine strain of Anaplasma centrale used in Australia. A monoclonal antibody that reacted with an 80 kDa antigen was used to develop an A. centrale-specific fluorescent antibody test that will be useful for confirming species identity in patent infections. Another monoclonal antibody that reacted with a 116 kDa antigen was used to develop an A. centrale-specific competitive inhibition enzyme-linked immunosorbent assay (ELISA) for the serological identification of vaccinated cattle. The sensitivity of the ELISA was 100% in cattle experimentally infected with A. centrale, 97.1% in a vaccinated beef herd and 98.3% in a vaccinated dairy herd. The specificity of the ELISA was 98.6% in non-vaccinated cattle outside the Anaplasma marginale-endemic area, 97.9% in non-vaccinated cattle within the A. marginale-endemic area and 100% in cattle experimentally infected with A. marginale. The ELISA detected antibodies to A. centrale in cattle up to 9 years after vaccination with no apparent decrease in sensitivity. The assay has proved extremely valuable in Australia for investigating reported failures of multivalent live vaccines used to protect cattle against anaplasmosis and babesiosis, and should be similarly useful elsewhere in the world where these types of vaccines are used, e.g. Israel and South America.

Anaplasma↗
PubMed2001

Major surface protein 1a effects tick infection and transmission of Anaplasma marginale.

Anaplasma marginale, an ehrlichial pathogen of cattle and wild ruminants, is transmitted biologically by ticks. A developmental cycle of A. marginale occurs in a tick that begins in gut cells followed by infection of salivary glands, which are the site of transmission to cattle. Geographic isolates of A. marginale vary in their ability to be transmitted by ticks. In these experiments we studied transmission of two recent field isolates of A. marginale, an Oklahoma isolate from Wetumka, OK, and a Florida isolate from Okeechobee, FL, by two populations of Dermacentor variabilis males obtained from the same regions. The Florida and Oklahoma tick populations transmitted the Oklahoma isolate, while both tick populations failed to transmit the Florida isolate. Gut and salivary gland infections of A. marginale, as determined by quantitative PCR and microscopy, were detected in ticks exposed to the Oklahoma isolate, while these tissues were not infected in ticks exposed to the Florida isolate. An adhesion-recovery assay was used to study adhesion of the A. marginale major surface protein (MSP) 1a to gut cells from both tick populations and cultured tick cells. We demonstrated that recombinant Escherichia coli expressing Oklahoma MSP1a adhered to cultured and native D. variabilis gut cells, while recombinant E. coli expressing the Florida MSP1a were not adherent to either tick cell population. The MSP1a of the Florida isolate of A. marginale, therefore, was unable to mediate attachment to tick gut cells, thus inhibiting salivary gland infection and transmission to cattle. This is the first report of MSP1a being responsible for effecting infection and transmission of A. marginale by Dermacentor spp. ticks. The mechanism of tick infection and transmission of A. marginale is important in formulating control strategies and development of improved vaccines for anaplasmosis.

Anaplasma↗