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PubMed2005

Comparison of three oxytetracycline regimes for the treatment of persistent Anaplasma marginale infections in beef cattle.

Anaplasmosis, caused by the tick-borne rickettsia, Anaplasma marginale, is an economically important disease of cattle in the United States and worldwide. Cattle that recover from acute infection become carriers in which low or microscopically undetectable A. marginale rickettsemia persists. Tetracycline antimicrobials are currently the only drug used in the US for treatment of acute anaplasmosis. There are currently no drugs specifically licensed for elimination of persistent infections. This study tested the efficacy of three oxytetracycline treatment regimens to clear A. marginale from cattle that were persistently infected. Forty Angus x Simmental steers, aged 6-12 months were experimentally infected with A. marginale. After the steers recovered from acute infection, seroconverted, and were confirmed infected using nested PCR followed by DNA hybridization, the carrier status of each animal was ascertained by sub-inoculation of blood into a separate, splenectomized Holstein calf. The steers were then blocked by bodyweight and randomly assigned as follows to four treatment groups: Treatment A, 300 mg/ml solution of oxytetracycline (Tetradure LA-300, Merial Canada Inc.) administered at 30 mg/kg, by intramuscular (i.m.) injection on day 0; Treatment B, the same 300 mg/ml solution of oxytetracycline administered at 30 mg/kg, i.m. on day 0 and again on day 5; Treatment C, a 200 mg/ml solution of oxytetracycline (Liquamycin LA-200, Pfizer Animal Health) administered at 22 mg/kg, intravenously (i.v.), q 24 h for 5 days (a treatment dose that corresponds with current Office International des Epizooties (OIE) recommendations for treatment prior to export). The fourth group consisted of untreated infected control cattle. All steers were still nested PCR and cELISA positive at 60 days after treatment. Infection was confirmed by subinoculation of blood into a splenectomized Holstein calf. These results demonstrated that the treatment regimens tested failed to clear A. marginale infections in carrier cattle.

Anaplasma marginale↗

PubMed2006

Prevalence and incidence of tick-borne diseases in smallholder farming systems in the western-Kenya highlands.

The prevalence and incidences of tick-borne diseases (TBDs) infections in cattle were studied in the western-Kenya highlands. Serological tests, thick-blood and lymph-node smears were used to quantify TBDs during cross-sectional and longitudinal studies. Four hundred and eight and 192 (wet season) and 114 and 46 (dry season) cattle were bled for serology in the rural and peri-urban areas, respectively. Seroprevalences differed significantly between the two areas (p<0.05). The cattle were monitored for 23 months (259,923 and 126,273 cattle days in the rural and peri-urban areas, respectively) from September 1996 to August 1998. The incidences of babesiosis, anaplasmosis and theileriosis were 0.42%, 4.64% and 4.92% and 1.45%, 32.11% and 39.05% in the rural and peri-urban areas, respectively. The difference in the incidence were significantly different for anaplasmosis and theileriosis (p<0.05). The difference in disease frequencies in two areas in close proximity shows the importance of monitoring disease events in order to understand and advice farmers in different production and farming systems appropriately.

Anaplasma↗

PubMed2000

Anaplasma marginale inactivated vaccine: dose titration against a homologous challenge.

The present study was performed to dose-titrate an Anaplasma marginale experimental immunogen derived from partially purified initial bodies from three geographically different Mexican strains. Three five-bovine groups were inoculated twice on days zero and 21 with A. marginale initial bodies equivalent to 1.5 x 10(10) (group I), 3 x 10(10) (group II) or 6 x 10(10) (group III) infected erythrocytes mixed with STDCM adjuvant. A similar group served as non-vaccinated controls. All four groups were challenged with 1 x 10(8) infected erythrocytes from a donor cow with an increasing rickettsemia of strain MEX-15 on day 87 post-vaccination. The prepatent period was very similar for all four groups. All five non-vaccinated controls presented typical acute anaplasmosis syndrome reaching a mean of 30.9% rickettsemia and a loss of 73.4% in the packed cell volume (PCV). Two of five controls died of acute anaplasmosis. Within the vaccinated groups only one animal (group II) suffered acute disease and died. Although all the other vaccinated animals were free of clinical signs, they developed very low rickettsemias (3.2, 3.8 and 4.3%) and PCV losses of 49.9, 47.8, and 49.3% for groups I, II and III. The starting mean weight was very similar for all four groups. All animals lost weight following challenge but losses for groups I and II were lower and significantly different from group IV losses (P < or = 0.1). Although there were no significant differences among vaccinated groups, group III was more severely affected. Taken altogether, these results show a 93.3% protection against both illness and death for all groups; and 100% protection for groups I and III, and 80% for group II.

Adjuvants, Immunologic↗

PubMed2001

Molecular phylogeny and biogeography of North American isolates of Anaplasma marginale (Rickettsiaceae: Ehrlichieae).

Anaplasma marginale (A. marginale) is a tick-borne ehrlichial pathogen of cattle that causes the disease anaplasmosis. Six major surface proteins (MSPs) have been identified on A. marginale from cattle and ticks of which three, MSP1a, MSP4 and MSP5, are from single genes and do not vary within isolates. The other three, MSP1b, MSP2 and MSP3, are from multigene families and may vary antigenically in persistently infected cattle. Several geographic isolates have been identified in the United States which differ in morphology, protein sequence and antigenic properties. An identifying characteristic of A. marginale isolates is the molecular weight of MSP1a which varies in size among isolates due to different numbers of tandemly repeated 28-29 amino acid peptides. For these studies, genes coding for A. marginale MSP1a and MSP4, msp1alpha and msp4, respectively, from nine North American isolates were sequenced for phylogenetic analysis. The phylogenetic analysis strongly supports the existence of a south-eastern clade of A. marginale comprised of Virginia and Florida isolates. Analysis of 16S rDNA fragment sequences from the A. marginale tick vector, Dermacentor variabilis, from various areas of the United States was used to evaluate possible vector-parasite co-evolution. Our phylogenetic analysis supports identity between the most parsimonious tree from the A. marginale MSP gene data and the tree that reflected the western and eastern clades of D. variabilis. These phylogenetic analyses provide information that may be important to consider when developing control strategies for anaplasmosis in the United States.

Amino Acid Sequence↗

PubMed2000

Establishment and characterization of an Oklahoma isolate of Anaplasma marginale in cultured Ixodes scapularis cells.

Anaplasma marginale is a tick-borne hemoparasite of cattle worldwide. The Virginia isolate of A. marginale was propagated previously in a cell line derived from embryos of the tick, Ixodes scapularis. The cultured Anaplasma (VA-tc) was passaged continuously for over 4 years and retained its infectivity for cattle and antigenic stability. We report herein the continuous in vitro cultivation of a second isolate of A. marginale derived from a naturally infected cow in Oklahoma (OK-tc). Blood from the infected cow was subinoculated into a splenectomized calf and blood collected at peak parasitemia was frozen, thawed and used as inoculum on confluent tick cell monolayers. Colonies of Anaplasma were apparent in low numbers at 9 days post exposure (PE) and infection in monolayers reached 100% by 4-5 weeks PE. Cultures were passaged by placing supernatant onto fresh tick cell monolayers at a dilution of 1:5 or 1:10. By the third passage development of the OK-tc was similar to that of the VA-tc and a 1:5 dilution resulted in 100% infection in 10-12 days. Inoculation of OK-tc into a splenectomized calf caused clinical anaplasmosis and Dermacentor ticks that fed on this calf transmitted the organism to a second susceptible calf. Major surface proteins (MSPs) 1-5 of the OK-tc were compared with homologous proteins present on VA-tc and the erythrocytic stage of the Oklahoma isolate. The MSPs 1, 2, 4, 5 were conserved on the OK-tc but there was evidence for structural variation in MSP3 between the cultured and erythrocytic stage of Anaplasma. MSP2 and MSP3 were the major proteins recognized by serum from infected cattle. Two-dimensional gels also identified positional differences between VA-tc and OK-tc in MSP2 and MSP3. The OK-tc may have potential to be used as antigen for development of an improved vaccine for anaplasmosis in the South Central United States.

Anaplasma↗

PubMed2002

A msp1alpha polymerase chain reaction assay for specific detection and differentiation of Anaplasma marginale isolates.

Anaplasma marginale is the causative agent of bovine anaplasmosis, a disease which can be protected by vaccination with the less pathogenic Anaplasma species, A. centrale. Currently, there is no polymerase chain reaction (PCR) assay available which differentiates between different species of Anaplasma or which can differentiate isolates of A. marginale within outbreaks and between different countries. A molecular test specific for A. marginale would be ideal for the identification of Anaplasma species in wild ruminants, as possible reservoirs of anaplasmosis, and to differentiate between A. marginale from A. centrale. A PCR assay was designed to amplify the major surface protein 1alpha gene of the rickettsial bovine pathogen, A. marginale both as an inter- and intra-specific test. The test did not amplify A. centrale or A. ovis, and discriminated A. marginale by amplifying repeat regions within the msp1alpha gene which vary in number between many isolates. The nested A. marginale amplicons varied in size from 630 to 1190bp representing one to eight internal repeats. All 22 Australian isolates tested amplified a 630bp product (one repeat) in contrast to all 19 non-Australian isolates tested. Eight sequences from Australian isolates from different geographical regions confirmed the conserved nature of the Australian A. marginale msp1alpha genes. The Australian 'repeat unit' MSP1a deduced amino acid sequence has been designated as Australian type 1. The msp1alpha PCR method developed here enabled the amplification and comparison of A. marginale isolates originating from North and South America, Africa, Israel and Australia. The method is sensitive and specific for A. marginale. Although additional msp1alpha products were amplified from at least two Australian isolates, the results suggest limited introduction of A. marginale into Australia.

Amino Acid Sequence↗

PubMed2004

Anaplasma phagocytophilum ligation to toll-like receptor (TLR) 2, but not to TLR4, activates macrophages for nuclear factor-kappa B nuclear translocation.

Anaplasma phagocytophilum is an obligate intracellular bacterium that infects neutrophils and causes human anaplasmosis (formerly human granulocytic ehrlichiosis). Interferon (IFN)- gamma causes immunopathology in A. phagocytophilum infection models. Plasma IFN- gamma levels are elevated 4 h after infection in experimentally infected mice, which indicates innate immunity and possible Toll-like receptors (TLRs). The ability of A. phagocytophilum to trigger proinflammatory responses via nuclear factor (NF)- kappa B was tested in TLR2- and TLR4-transfected cell lines and in primary murine macrophages devoid of TLR2 or TLR4. NF- kappa B was activated only through TLR2, which suggests its role in innate immune induction with A. phagocytophilum infections. The role of innate immunity in human anaplasmosis immunopathology requires more study.

Anaplasma phagocytophilum↗

PubMed1992

Seroprevalence survey for Anaplasma marginale-infection of Austrian cattle.

A serologic survey study of 5,076 Austrian cattle farming herds was carried out in the period of December 1988 till March 1990. One animal was randomly selected from each herd and the antibody titer against Anaplasma marginale in blood serum samples was evaluated by means of the complement fixation test. The number of these tested blood samples was 3.6% of 140,081 cattle herd farms of Austria. 109 (2.1%) of the tested animals showed positive titers (1:10) against Anaplasma marginale, in relation to the 140,081 cattle herds 0.08%, 4,786 (94.3%) blood serum samples were sero-negative, 188 (3.8%) reacted anticomplementary. The highest number of antibody-positive animals of 8 tested Austrian districts could be found in Carinthia (46 = 5.7%). In Burgenland all tested sera turned out to be negative. Concerning the distribution of sero-positive animals in Austria it can be stated that a decrease of positive reactors from southern to northern region is evident. A connection between the occurrence of anaplasmosis in Italy, Yugoslavia, Switzerland and Hungary, is postulated as a result of the different systems of keeping cattle in the provinces and the regional increase of tick invasion. Possibly an intensive animal transportation is of importance due to the introduction of the disease mentioned before. The results obtained show that anaplasmosis does occur in different areas of Austria. For control of this disease in Austria it is proposed that all imported cattle should be tested serologically for antibodies against Anaplasma marginale. Other diseases in connection with anemia should be excluded by clinical, serological, blood-, as well as pathological examinations.

Anaplasma↗

PubMed2005

Genetic diversity of Anaplasma marginale strains from cattle farms in the province of Palermo, Sicily.

Bovine anaplasmosis, caused by the tick-borne rickettsia Anaplasma marginale, is endemic in Sicily and results in economic loss to the cattle industry. This study was designed to characterize strains of A. marginale at the molecular level from cattle in the Province of Palermo, Sicily. Seropositivity of cattle >or=1 year old for A. marginale in the study area ranged from 62% to 100%. The observed prevalence of A. marginale infections in cattle herds ranged from 25% to 100%. Two predominant A. marginale msp4 genotypes were found. A positive correlation was found between the prevalence of infection and the presence of Rhipicephalus (Boophilus) annulatus. Phylogenetic analysis of msp4 sequences of European strains of A. marginale did not provide phylogeographical information. These results suggest that development of farm husbandry systems and vaccines for genetically heterogeneous populations of A. marginale are needed for control of anaplasmosis in this region of Sicily.

Anaplasma marginale↗

PubMed2006

Phylogenetic analysis of Anaplasma marginale strains from Paraná State, Brazil, using the msp1alpha and msp4 genes.

Anaplasma marginale is an obligate intraerythrocytic rickettsial pathogen (order, Rickettsiales: family, Anaplasmataceae) that causes bovine anaplasmosis. This disease is widely distributed in tropical and sub-tropical regions of the world and causes important economic losses to cattle production. Major surface protein (MSP)1a (msp1alpha gene) is one of the six MSPs identified on A. marginale from cattle, whose sequence and size vary according to the number of tandem 28- to 29-amino acid repeats. This study characterized the msp1alpha and msp4 genes obtained from three distinct Brazilian herds from the State of Paraná. Three strains of the msp1alpha and one strain of the msp4 gene were sequenced. The strains evaluated revealed PCR products of different size, representing three, five and six internal repeats. Sequence analyses confirmed the number of tandem sequence copies and revealed a high degree of sequence identity with strains from other Brazilian States, as well as strains from the USA, Europe and Israel. The msp1alpha DNA and amino acid sequences from A. marginale and DNA sequences of msp4 strains did not reveal distinct phylogeographical segregation. However, the amino acid sequences of msp4 demonstrated definite phylogeographical relationship. These results suggest that the amino acid sequences of msp4 should be used for phylogenetic identification of A. marginale strains and may be an important tool for the epidemiology and control of anaplasmosis. Additionally, the close similarity of the Paraná strains of A. marginale with strains from USA, Europe and Asia may reflect the introduction of these genes during the development of the Brazilian bovine herd.

Amino Acid Sequence↗

PubMed1996

Effect on intraerythrocytic Anaplasma marginale of soluble factors from infected calf blood mononuclear cells.

Blood mononuclear cells (lymphocytes and monocytes) were isolated from infected calves during in vivo control of acute anaplasmosis and cultured with Anaplasma marginale organisms. Supernatants from the cultures reduced the proportion of erythrocytes containing viable A. marginale in vitro, indicating that an antibody-independent mechanism of rickettsemia control might occur during acute anaplasmosis.

Acute Disease↗

PubMed1991

Identification of Anaplasma marginale long-term carrier cattle by detection of serum antibody to isolated MSP-3.

Rapid and accurate detection of Anaplasma marginale-infected cattle would enhance anaplasmosis control procedures and evaluation of vaccines. Current tests based on detection of antibodies in serum are not widely used for several reasons, including the occurrence of either false-positive or false-negative results. We evaluated binding of antibodies in serum to a subunit antigen isolated from A. marginale initial bodies--major surface protein 3 (MSP-3). MSP-3 was detected in lysates of eight geographically different isolates of A. marginale and purified by affinity chromatography with monoclonal antibody AmG75C2. Antibodies from cattle infected with any of five geographically different isolates of A. marginale reacted in immunoblots with MSP-3. Sera from uninfected cattle and cattle infected with another rickettsial organism and two hemoprotozoal organisms failed to react with MSP-3. Six carrier cattle infected with the Florida isolate of A. marginale had antibody titers to MSP-3 ranging from 10(3) to 10(6) during a 5-year evaluation period. Since specific antibodies to isolated MSP-3 persist in high titers in long-term carrier cattle sera and MSP-3 is common among A. marginale isolates, it is recommended as a subunit antigen for an anaplasmosis test.

Anaplasma↗

PubMed1975

Performance of Holstein and crossbred dairy cattle in Louisiana. III. Health and viability.

Viability and health disorders were compared for straightbred Holsteins and crosses between Holsteins, Brown Swiss, Jerseys, and Red Sindhis. In addition, effects of mating systems and effects of various parts of Red Sindhi heredity on these traits were evaluated. Losses before first calving per 100 females born alive were: Holsteins 22; Holstein-sired crosses 20: Brown Swiss-sired crosses 26; daughters of crossbred sires group I (sires had from 1/4 to 1/16 Red Sindhi heredity) 24; and group II (sires were all European breeds) 31. Losses in first-gestation from abortions and stillbirths were: Holsteins 20% and crossbreds 9%. In first lactation, incidence of foot rot and anaplasmosis was higher among Holsteins (13% and 16%) than in crossbreds (5% and 8%) whereas the incidence was less among crosses with Red Sindhi heredity (3% and 2%) than in all European breed crosses (9% and 17%). When progeny produced by criss-crossing to purebred sires were compared with progeny from random mating to crossbred sires, the former had a greater incidence of mastitis (30% versus 19%) and anaplasmosis (11% versus 2%). There was no differences between Holsteins and crossbreds in overall viability. However, there was considerable variation between breeds at the various ages.

Abortion, Veterinary↗

PubMed1987

DNA probes for the detection of Anaplasma centrale and Anaplasma marginale.

Anaplasmosis can be diagnosed either by immunological techniques or by direct microscopic examination of blood smears. Both methods are time-consuming and labour intensive. The use of DNA probes in an hybridization assay may simplify the diagnosis of anaplasmosis in cattle and sheep. A genomic DNA library of Anaplasma centrale was constructed in an expression vector and screened to detect clones containing A. centrale DNA. Four probes which hybridized to A. centrale and Anaplasma marginale DNA were isolated. One of these (AC-1) hybridized only to A. centrale DNA, whereas AC-2, AC-3 and AC-4 could detect DNA from both A. centrale and A. marginale. Probes AC-1 and AC-2 could detect 127 ng and 8 ng DNA respectively, while AC-3 and AC-4 detected 64 ng A. centrale DNA.

Anaplasma↗

PubMed1987

Attempted transmission to cattle of Anaplasma marginale from overwintered Dermacentor andersoni ticks.

Since the 1983 summer outbreak of anaplasmosis in southern Saskatchewan, the role of the tick, Dermacentor andersoni as an overwintering reservoir for Anaplasma marginale has been questioned. The purpose of this study was to determine if spring-collected ticks carried virulent A. marginale. Sixteen splenectomized calves were assigned randomly to two groups of 14 principals and two controls. Adult D. andersoni, collected in April from areas having high transmission rates of A. marginale, were confined to the ears of the principals by special bags and allowed to feed for eight days. The two control calves were subsequently challenged intravenously with blood from a calf infected with the Virginia strain of A. marginale. Principals and controls were monitored for 60 and 50 days postexposure respectively for signs of infection by clinical, hematological and serological procedures. None of the principals developed anaplasmosis but both control calves developed signs of disease.

Anaplasmosis↗

PubMed1986

Transmission of Anaplasma marginale Theiler by males of Dermacentor andersoni Stiles fed on an Idaho field-infected, chronic carrier cow.

The role of ticks and carrier cattle in epizootics of bovine anaplasmosis was further clarified by demonstrating unequivocally, for the first time, that male ticks fed on a chronic carrier cow naturally infected with Anaplasma marginale can transmit this parasite intrastadially and biologically when subsequently fed on susceptible cattle. These data indicate that field epizootics of acute anaplasmosis may be initiated by males of tick vector species that feed on carrier cattle and subsequently transfer to susceptible cattle.

Anaplasma↗

PubMed1978

Serologic and hematologic response of rabbits to Anaplasma marginale-infected bovine erythrocytes.

New Zeland White rabbits inoculated with Anaplasma marginale-infected bovine erythrocytes produced specific humoral antibodies to this parasite. Specific antibody production as measured by the card and complement-fixation (CF) tests was maximal in 7 to 21 days after inoculation; however, none of the rabbits became infected. Geometric mean CF titers of rabbits inoculated with RBC from cattle with acute anaplasmosis were significantly higher (P less than 0.001) than those of rabbits given RBC from normal cattle. Significant difference was not seen between geometric mean CF titers of rabbits given RBC from carrier cattle and those of rabbits given RBC from normal cattle. It is concluded, therefore, that rabbit inoculations cannot effectively be substituted for calf inoculations to determine the anaplasmosis status of carrier cattle.

Agglutination Tests↗

PubMed1984

Immune responses of calves antigenically stimulated and challenge exposed with Anaplasma marginale during tick infestation or treatment with dexamethasone.

Similar anamnestic antibody responses to a 2nd injection of a Anaplasma marginale vaccinal antigen were observed in calves infested with the tick Dermacentor albipictus and in tick-free calves. When challenge exposure of these calves to virulent A marginale was done, infestation with the tick Boophilus microplus increased anemia (P less than 0.01), but did not suppress antibody production to A marginale or increase parasitemia. None of the vaccinated calves, regardless of infestation, experienced clinical anaplasmosis. Mitogenic responses of lymphocytes from infested animals were unaltered by either infestation. Tick infestations did not cause immune suppression in the calves, whereas use of dexamethasone resulted in a significantly lower antibody response (P less than 0.05) after a 2nd injection of vaccinal antigen. After challenge with virulent A marginale, dexamethasone-treated calves showed more pronounced parasitemia (P less than 0.01) and anemia (P less than 0.01) than did control calves. Anaplasmosis did not prevent the calves from developing resistance to reinfestation, which was accompanied by immediate, but not delayed, hypersensitivity reactions against homologous tick extracts. Dermacentor albipictus did not seem to share common antigenic determinants with B microplus, since extracts from the latter did not elicit immediate hypersensitivity responses in calves sensitive to D albipictus extracts. Calves were somewhat resistant to reinfestation as evidenced by reduced numbers of adult fed ticks, decreased weights of ticks after feeding, and smaller egg masses.

Anaplasmosis↗