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An epidemiological study on Anaplasma infection in cattle, sheep, and goats in Mashhad Suburb, Khorasan Province, Iran.

The prevalence of Anaplasma infection was studied in cattle, sheep, and goats in the Mashhad area from 1999 to 2002. A total of 160 cattle from 32 farms and 391 sheep and 385 goats from 77 flocks were clinically examined for the presence of Anaplasma spp. in blood smears. The study revealed that 19.37% of cattle were infected with Anaplasma marginale and 80.3% of sheep and 38.92% of goats were infected with Anaplasma ovis. Prevalence of Anaplasma infection between male and female and between different age groups of cattle, sheep, and goats were statistically nonsignificant. Seasonally, the prevalence of Anaplasma infection in sheep and goats reached its highest level in summer, while a decrease was observed in autumn, and reached the lowest level in winter. The seasonal prevalence of Anaplasma infection in cattle was not significantly different. Symptomatic cases were not observed in any of the cattle, sheep, and goats. The ranges of anaplasmatemia in infected cattle, sheep, and goats were 0.005-0.5%, 0.01-3%, and 0.01-3%, respectively.

Anaplasmosis↗

Identification of Borrelia burgdorferi sensu lato, Anaplasma and Ehrlichia species, and spotted fever group Rickettsiae in ticks from Southeastern Europe.

Prevalence data for tick-borne pathogens are used to assess the risk for human health. In this study the presence and identity of Borrelia burgdorferi sensu lato, Ehrlichia, Anaplasma, and Rickettsia species in Bulgarian Ixodes ricinus ticks and in non- Ixodes ticks from Turkey and Albania was determined by polymerase chain reaction (PCR) and reverse line blot hybridization. In the adult Bulgarian ticks, the prevalence of Borrelia burgdorferi sensu lato infection was approximately 40%, while Borrelia afzelii was the predominant species, representing more than half of all Borrelia-positive ticks. Ehrlichia and Anaplasma species were detected in 35% of the adult Ixodes ricinus ticks and in 10% of the nymphs. Sequence analysis of PCR products reacting with the Anaplasma phagocytophila probe revealed a 16S rRNA gene identical to that of the Anaplasma phagocytophila prototype strain. Ehrlichia and Anaplasma species were found in approximately 7% of the non- Ixodes ticks. Sequence analysis of some of these samples revealed the presence of Anaplasma ovis, Ehrlichia canis, and a species closely resembling Ehrlichia chaffeensis. About half of all adult ticks examined and approximately 20% of all nymphs were infected with Rickettsia species. In Ixodes ricinus ticks, Rickettsia helvetica and a Rickettsia species designated as IRS3 were found in high prevalence. Rickettsia conorii was found in virtually all non- Ixodes tick species from Albania and Turkey. The results of this study show that many tick-borne diseases are most probably endemic in the Balkan area. Furthermore, the results suggest that there is a considerable chance for simultaneous transmission of tick-borne pathogens to human beings.

Albania↗

Humoral immune response and hematologic evaluation of pregnant Jersey cows after vaccination with Anaplasma centrale.

The main objective of this work was to evaluate the safety of an Anaplasma centrale vaccine in pregnant pure bred Jersey cows selected from a herd located at Miranda State, Venezuela. Ten cows of 3-5 months of gestation were chosen and previous vaccination all cows were tested for Anaplasma antibodies by the indirect immunofluorescence assay (IFA), so only seronegative cows were included in the group, and for blood parameters, rectal temperature, and pregnancy. Selected cows were vaccinated intramuscularly with 1ml of an A. centrale live vaccine which had 10(8) A. centrale per ml. Over the next 2 months cows were checked weekly for hematological parameters and Anaplasma antibodies, and then for the next 2 months these evaluations were performed monthly. Among the values monitored were: A. centrale parasitemia, hematocrit, hemoglobin, and white blood cells (WBCs) (neutrophil, lymphocyte and eosinophil counts). Levels of Anaplasma antibodies were measured by IFA. Anaplasma were observed for the first time in blood films of two vaccinated cows at 14 days post-vaccination (PV), 6 out of 10 cows were A. centrale positive at 30 days PV, and all cows were A. centrale positive at 42 days PV. A. centrale often showed low parasitemia, 1-3%. Anaplasma antibodies were detected at day 14 PV in all vaccinated cows with a mean group titre of 360 (range: 80-1280). All vaccinated cows showed few changes in their hematologic parameters or in rectal temperature, and all gave birth to healthy calves. In conclusion, adult pregnant cows were safely vaccinated with this live A. centrale vaccine, which may help to develop a cross-protective immunity against field strains of A. marginale.

Anaplasma↗

Detection of Ehrlichia spp., Anaplasma spp., Rickettsia spp., and other eubacteria in ticks from the Thai-Myanmar border and Vietnam.

A total of 650 ticks, including 13 species from five genera, were collected from animals, from people, or by flagging of the vegetation at sites on the Thai-Myanmar border and in Vietnam. They were tested by PCR to detect DNA of bacteria of the order RICKETTSIALES: Three Anaplasma spp. were detected in ticks collected in Thailand, including (i) Anaplasma sp. strain AnDa465, which was considered a genotype of Anaplasma platys (formerly Ehrlichia platys) and which was obtained from Dermacentor auratus ticks collected from dogs; (ii) Anaplasma sp. strain AnAj360, which was obtained from Amblyomma javanense ticks collected on a pangolin; and (iii) Anaplasma sp. strain AnHl446, which was closely related to Anaplasma bovis and which was detected in Haemaphysalis lagrangei ticks collected from a bear. Three Ehrlichia spp. were identified, including (i) Ehrlichia sp. strain EBm52, which was obtained from Boophilus microplus ticks collected from cattle from Thailand; (ii) Ehrlichia sp. strain EHh324, which was closely related to Ehrlichia chaffeensis and which was detected in Haemaphysalis hystricis ticks collected from wild pigs in Vietnam; and (iii) Ehrlichia sp. strain EHh317, which was closely related to Ehrlichia sp. strain EBm52 and which was also detected in H. hystricis ticks collected from wild pigs in Vietnam. Two Rickettsia spp. were detected in Thailand, including (i) Rickettsia sp. strain RDla420, which was detected in Dermacentor auratus ticks collected from a bear, and (ii) Rickettsia sp. strain RDla440, which was identified from two pools of Dermacentor larvae collected from a wild pig nest. Finally, two bacteria named Eubacterium sp. strain Hw124 and Eubacterium sp. strain Hw191 were identified in Haemaphysalis wellingtoni ticks collected from chicken in Thailand; these strains could belong to a new group of bacteria.

Anaplasma↗

Isolation of an Anaplasma sp. organism from white-tailed deer by tick cell culture.

We used tick cell culture to isolate a bacterium previously referred to as the "white-tailed deer (WTD) agent" from two captive fawns inoculated with blood from wild WTD (Odocoileus virginianus). Buffy coat cells were added to ISE6 tick cell cultures and incubated at 34 degrees C, and 8 days later, Anaplasma-like inclusions were demonstrated in Giemsa-stained culture samples. The microbes became established and could be continuously passaged in tick cells. The identity of a culture isolate designated WTD76 was verified as the WTD agent by using specific PCR primers and by DNA sequencing. Comparison with sequences available in GenBank indicated that the isolate was most closely related first to Anaplasma platys and second to Anaplasma phagocytophilum, supporting its placement in the genus Anaplasma. Transmission electron microscopy of this Anaplasma sp. organism in tick cell cultures revealed large inclusions filled with pleomorphic and rod-shaped bacteria. Tick cells infected with the Anaplasma sp. organism were used to successfully infect a naive deer, thereby proving the infectivity of the isolate for deer.

Anaplasma↗

Amplification of 16S rRNA genes of Anaplasma species in China for phylogenetic analysis.

In this study, a phylogenetic tree was inferred through comparing five 16S rRNA gene sequences of four isolates of Anaplasma ovis and one of Anaplasma marginale in China with all nineteen 16S rRNA gene sequences deposited in GenBank (12 A. marginale, 3 A. ovis and 4 Anaplasma centrale derived from America, Uruguay, South Africa, Zimbabwe, Australia, Isreal and Japan). The analysis showed that all A. ovis isolated in China were separated into an A. ovis cluster, while the A. marginale in China was separated into an A. marginale cluster (see Fig. 1). This analysis demonstrated that there are at least two different Anaplasma species widespread among ruminants in North China.

Anaplasma↗

Antigen recognition on Anaplasma marginale and bovine erythrocytes: an electron microscopy study.

The reactivity of sera from Anaplasma marginale-infected bovine with red blood cells and with purified anaplasma bodies was analyzed by electron immunomicroscopy. Red blood cells from non-infected and from anaplasma-infected cows and A. marginale bodies separated from parasitized erythrocytes, were incubated with control, pre-immune and immune sera followed by anti-bovine IgG-Peroxidase. Immune sera from cows infected with the venezuelan and Florida isolate reacted with red blood cell membranes from normal and infected bovines, while sera from non-infected cows did not. The immune sera also recognized epitopes localized on the cell wall, membrane and on unidentified intracellular structures of the purified anaplasma bodies. Thus, we propose that A. marginale infection may cause structural and biochemical modifications of the plasma membrane of the bovine red blood cells during its intraerythrocytic cycle. This in turn could elicit an autoimmune type of response against its own cells that would stimulate erythrophagocitosis. The strong reactivity of the immune sera with the Anaplasma bodies suggests that the bovine immune system also recognizes epitopes located on the parasite.

Anaplasma↗

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↗

Serological survey of Ehrlichia and Anaplasma infection of feral raccoons (Procyon lotor) in Kanagawa Prefecture, Japan.

Numbers of feral raccoon; the possible reservoir animal of Ehrlichia and Anaplasma, are increasing in Japan. Thus serological methods were utilized to examine Ehrlichia and Anaplasma infection in raccoons from Kanagawa Prefecture, Japan. By using an indirect immunofluorescence assay, among 187 feral raccoons examined, 1 (0.5%) serologically reacted with Ehrlichia canis, 3 (1.6%) with Ehrlichia chaffeensis and 1 (0.5%) with Anaplasma phagocytophilum with the titers of 1:40 or more. Although screening PCR for Ehrlichia and Anaplasma species failed to detect the presence of ehrlichial DNA in serum samples, results of the serological tests suggested that the feral raccoons might be infected with some species of Ehrlichia and Anaplasma.

Anaplasma↗

Simultaneous detection of Anaplasma and Ehrlichia species in ruminants and detection of Ehrlichia ruminantium in Amblyomma variegatum ticks by reverse line blot hybridization.

The detection of Anaplasma and Ehrlichia species is usually based on species-specific PCR assays, since no assay is yet available which can detect and identify these species simultaneously. To this end, we developed a reverse line blot (RLB) assay for simultaneous detection and identification of Anaplasma and Ehrlichia species in domestic ruminants and ticks. In a PCR the hypervariable V1 region of the 16S ribosomal RNA (rRNA) gene was amplified with a set of primers unique for members of the genera Anaplasma and Ehrlichia [Int. J. Syst. Evol. Microbiol. 51 (2001) 2145]. Amplified PCR products from blood of domestic ruminants or Amblyomma variegatum tick samples were hybridized onto a membrane to which eight species-specific oligonucleotide probes and one Ehrlichia and Anaplasma catch-all oligonucleotide probe were covalently linked. No DNA was amplified from uninfected blood, nor from other hemoparasites such as Theileria annulata, or Babesia bigemina. The species-specific probes did not cross-react with DNA amplified from other species. E. ruminantium, A. ovis and another Ehrlichia were identified by RLB in blood samples collected from small ruminants in Mozambique. Finally, A. variegatum ticks were tested after feeding on E. ruminantium infected sheep. E. ruminantium could be detected in adult ticks even if feeding of nymphs was carried out 3.5 years post-infection. In conclusion, the developed species-specific oligonucleotide probes used in an RLB assay can simultaneously detect and identify several Ehrlichia and Anaplasma species. However, as no quantitative data for the detection limit are available yet, only positive results are interpretable at this stage.

Anaplasma↗

Molecular epidemiological study for tick-borne disease (Ehrlichia and Anaplasma spp.) surveillance at selected U.S. military training sites/installations in Korea.

Vector-borne diseases are a potential public health threat to U.S. Forces Korea (USFK). Ehrlichia and Anaplasma spp., transmitted by ticks, are only two of several diseases that may affect military readiness and operations. Rodents were collected at selected U.S. military installations and training sites in the Republic of Korea. DNA was extracted from spleen tissues and assayed by PCR methods for Ehrlichia and Anaplasma species. From rodents and mustelids collected during 1999 and 2000, a total of 196 Apodemus agrarius (striped field mouse), 2 Mustela sibirica (weasel), and 1 Cricetulus triton nestor (Korean greater long-tailed hamster) were assayed for Ehrlichia and Anaplasma species-specific DNA fragments. Rodent surveillance indicated a very high prevalence of Ehrlichia and Anaplasma spp. at selected training sites. Ehrlichia/Anaplasma DNA were identified from spleen tissue from 157 Apodemus agrarius, 1 Mustela sibirica, and 1 Cricetulus riton nestor. Species-specific DNA fragments of E. canis (45), E. ewingii (16), A. phagocytophila (5), and A. platys (62) were amplified by PCR techniques. Seventy-one striped field mice had single infections, while 24 had mixed infections of 2 (17 specimens), 3 (7 specimens), or 4 (1 specimen) pathogens. The striped field mouse plays a role as a reservoir for latent infections of various Ehrlichia or Anaplasma species.

Anaplasma↗

Longitudinal analysis of tick densities and Borrelia, Anaplasma, and Ehrlichia infections of Ixodes ricinus ticks in different habitat areas in The Netherlands.

From 2000 to 2004, ticks were collected by dragging a blanket in four habitat areas in The Netherlands: dunes, heather, forest, and a city park. Tick densities were calculated, and infection with Borrelia burgdorferi and Anaplasma and Ehrlichia species was investigated by reverse line blot analysis. The lowest tick density was observed in the heather area (1 to 8/100 m2). In the oak forest and city park, the tick densities ranged from 26 to 45/100 m2. The highest tick density was found in the dune area (139 to 551/100 m2). The infection rates varied significantly for the four study areas and years, ranging from 0.8 to 11. 5% for Borrelia spp. and 1 to 16% for Ehrlichia or Anaplasma (Ehrlichia/Anaplasma) spp. Borrelia infection rates were highest in the dunes, followed by the forest, the city park, and heather area. In contrast, Ehrlichia/Anaplasma was found most often in the forest and less often in the city park. The following Borrelia species were found: Borrelia sensu lato strains not identified to the species level (2.5%), B. afzelii (2.5%), B. valaisiana (0.9%), B. burgdorferi sensu stricto (0.13%), and B. garinii (0.13%). For Ehrlichia/Anaplasma species, Ehrlichia and Anaplasma spp. not identified to the species level (2.5%), Anaplasma schotti variant (3.5%), Anaplasma phagocytophilum variant (0.3%), and Ehrlichia canis (0.19%) were found. E. canis is reported for the first time in ticks in The Netherlands in this study. Borrelia lusitaniae, Ehrlichia chaffeensis, and the human granylocytic anaplasmosis agent were not detected. About 1.6% of the ticks were infected with both Borrelia and Ehrlichia/Anaplasma, which was higher than the frequency predicted from the individual infection rates, suggesting hosts with multiple infections or a possible selective advantage of coinfection.

Anaplasma↗

Comparative study of Anaplasma parasites in tick carrying buffaloes and cattle.

A comparative study on the prevalence of Anaplasma parasite was conducted on ticks carrying buffaloes and cattle. Five hundred blood samples of both animals (250 of each) were collected during February, March and April. Thin blood smears on glass slides were made, fixed in 100% methyl alcohol and examined. Microscopic examination revealed that 205 (41%) animals had Anaplasma parasites, out of which 89, 44 and 72 animals had Anaplasma marginale, Anaplasma centrale and mixed infection respectively. Infected buffaloes and cattle were 75 and 130 respectively. The infection in female was 53 and 92 in buffaloes and cattle respectively. Twenty-two and 92 blood samples of male were found positive in buffaloes and cattle respectively. Comparative study revealed that the cattle were 26.82% more susceptible than buffaloes. The parasite prevailing percentage in female of both animals was slightly higher than that of the male. This investigation was aimed at studying the comparative prevalence of Anaplasma parasite in tick carrying buffaloes and cattle.

Anaplasma↗

Anaplasma infections in wild and domestic ruminants: a review.

Anaplasma marginale can be transmitted, will grow and can survive in a large number of domestic and wild animals. It is pathogenic in cattle, and usually produces nonapparent or mild infections in other species. Anaplasma marginale has been recovered from cattle, sheep, goats, water buffalo (Bubalus bubalis), white-tailed deer (Odocoileus virginianus), mule deer (Odocoileus hemionus hemionus), black-tailed deer (Odocoileus hemionus columbianus), pronghorn (Antilocapra americana americana), Rocky Mountain elk (Cervus elaphus nelsoni), bighorn sheep (Ovis canadensis canadensis), black wildebeest (Connochaetes gnu), blesbuck (Damaliscus albifrons), and duiker (Sylvicapra grimmi grimmi). Unidentified anaplasms have been seen in, and in some instances isolated from, Cape buffalo (Syncerus caffer), giraffe (Giraffa camelopardalis), wildebeest (Connochaetes taurinus), Cokes hartebeest (Alcelaphus buselaphus cokii), Thompson's gazelle (Gazella thompsonii), waterbuck (Kobus ellipsiprymnus), and sable antelope (Hippotragus niger), with serological evidence of Anaplasma infection in an even wider range of wild ruminant species. Anaplasma ovis, A. centrale, or other as yet unidentified anaplasms may well occur in other ruminants. With the exception of black-tailed deer, the epidemiologic significance of anaplasmosis in wildlife has yet to be determined. The only wild animal in which Anaplasma is reported to produce serious clinical disease is the giraffe.

Africa↗

Genetic characterization of Anaplasma ovis strains from bighorn sheep in Montana.

Wildlife reservoir species and genetic diversity of Anaplasma ovis (Rickettsiales: Anaplasmataceae) have been poorly characterized. Bighorn sheep (Ovis canadensis), captured in Montana from December 2004 to January 2005, were tested for antibodies to Anaplasma spp.; the presence of A. ovis was determined by the characterization of major surface protein msp4 sequences. Anaplasma antibodies were detected in 25/180 (14%) sampled bighorn sheep and A. ovis msp4 sequences were amplified by polymerase chain reaction (PCR) and sequenced from 9/23 (39%) of seropositive animals. All animals were negative by PCR for the related pathogens, Anaplasma phagocytophilum and Anaplasma marginale. All msp4 sequences identified in the bighorn sheep were identical and corresponded to a single A. ovis genotype that was identical to a sheep isolate reported previously from Idaho. The finding of a single genotype of A. ovis in this wild herd of bighorn sheep was in contrast to the genetic diversity reported for A. marginale in cattle herds in the western United States and worldwide. These results demonstrated that bighorn sheep may be a wildlife reservoir of A. ovis in Montana.

Anaplasma ovis↗

Morphologic alterations of Anaplasma marginale in calves after treatment with oxytetracycline.

The morphologic features of Anaplasma marginale were determined after treatment of infected calves with oxytetracycline. By light microscopy, anaplasma bodies in erythrocytes of treated calves were enlarged and vacuolated, small and dense, or comma shaped. Several degenerated forms of anaplasma bodies were observed by electron microscopy. There was a blending of the 2 membranes surrounding initial bodies, aggregation of nucleoprotein at the periphery of the subunit, and vacuolation. A 2nd form of degeneration was coalescence of subunits in marginal bodies. A 3rd form of degeneration was unification of subunits of anaplasma bodies and clumping of nucleoprotein. A 4th form of degeneration was the persistence of an anaplasma body as a solitary, irregularly shaped mass containing electron-opaque clumps.

Anaplasma↗

Efficacy of attenuated Anaplasma marginale vaccine under laboratory and field conditions in Colombia.

Four-month-old Holstein-Friesian calves were inoculated with 3 different doses (1, 2, and 3 ml) of attenuated Anaplasma marginale vaccine. Vaccinated calves showed mild anaplasma parasitemia, slight decrease in packed cell volume, low serologic conversion, and no clinical illness. An artificial challenge exposure of vaccinated and unvaccinated calves with virulent Colombian A marginale showed that the vaccine provided protection against clinical signs of the disease, including parasitemia and anemia. The volume of the vaccinal dose did not alter the degree of protection provided. A 2nd group of 8- to 9-month-old Holstein-Friesian calves was then inoculated with 3 ml of anaplasma vaccine and premunized with both Babesia bigemina and Babesia argentina while being housed in an area free of these diseases. Calves were moved to an enzootic region heavily infested with various arthropods, including ticks, for natural field challenge exposure. Control calves, which were not given anaplasma vaccine, suffered clinical illness manifested by severe anemia and an average weight loss of 50.6 kg due to anaplasma field challenge exposure. In contrast, vaccinated calves did not show anemia and their weight loss was 3.9 kg.

Anaplasma↗

The distribution of Dermacentor hunteri and Anaplasma sp. in desert bighorn sheep (Ovis canadensis).

The ixodid tick Dermacentor hunteri has been collected intermittently this century, primarily from desert bighorn sheep (Ovis canadensis). Anaplasma spp. are intraerythrocytic rickettsial parasites of ungulates and are vectored in the western United States by ticks of the genus Dermacentor. We tested the hypotheses that D. hunteri would be found infesting all populations of desert bighorn, and that all infested populations would be seropositive for Anaplasma sp. Dermacentor hunteri was found on desert bighorn throughout their range in the Mojave and Sonoran deserts of the southwestern United States and northern Mexico, but not in any portion of the Chihuahuan desert of New Mexico and eastern Arizona or in Baja California Sur, Mexico. Using an indirect immunofluorescence antibody test (IIF), 8 populations of desert bighorn in California with D. hunteri were seropositive for Anaplasma sp. (n = 160). Four populations of desert bighorn with D. hunteri in Arizona (n = 69), 1 in Nevada (n = 22), and I in Utah (n = 14) with D. hunteri were seronegative. Six populations of desert bighorn were uninfested with D. hunteri and were also seronegative. Of these populations, 1 was in California (n = 19), 2 were in New Mexico (n = 33), 2 were in Utah (n = 30), and 1 was in Baja California Sur (n = 14). We found no support for either of our original hypotheses and concluded that both D. hunteri and Anaplasma sp. are limited in their distribution among desert bighorn. We also suggest a cautionary approach to translocations of desert bighorn given the high prevalence of ticks and the unknown effects of Anaplasma sp. on free-ranging bighorn.

Anaplasma↗