Genetic, host and environmental factors associated with a high prevalence of Anaplasma marginale.

Anaplasma marginale is the most prevalent vector-borne pathogen in the livestock industry in Colombia, causing economic losses of approximately USD 4.2 million per year. The present study reports the seasonal transmission patterns, genetic diversity and phylogeographic traits of A. marginale strains in cattle and buffaloes from Colombian livestock areas. A three-point longitudinal survey was designed to evaluate the above characteristics of farms in the Caribbean and Orinoquía regions. The A. marginale prevalence was evaluated in 1432 cattle blood samples, 152 buffalo blood samples and the hemolymph of 439 ticks using semi-nested PCR (sn-PCR) targeting the msp5 gene. The molecular prevalence in cattle and buffaloes was 54.8% and 13.1%, respectively, with higher values during the wet and late wet seasons. Factors such as age and production system were significantly associated with the infection. Rhipicephalus microplus was the only carrier of A. marginale DNA, with an infection rate of 17.2%. On the other hand, the tandem repeat and microsatellite analyses of the msp1α gene showed high genetic diversity and new tandem repeats that suggested strain adaptation to different transmission modes. Phylogeographic analysis using the msp4 gene showed a relationship between Colombian isolates and Mexican, Brazilian, Venezuelan, European and Asian isolates, as well as two worldwide haplogroups that were associated with the geographical origin of each isolate. In conclusion, this study shows that A. marginale occurs under enzootic stability in both hosts, with a high prevalence of infection during wet months and in animals dedicated to beef production. The genetic variability analyses suggest that a high strain diversity is associated with multiple selective pressures in the study area, while phylogeographic traits suggest a high genetic similarity between Mexican and South American strains.

Cattle experimentally infected by Anaplasma marginale: Influence of splenectomy on disease pathogenesis, oxidative profile, and antioxidant status.

Bovine anaplasmosis is caused by the obligate intraerythrocytic bacteria Anaplasma marginale. These bacteria are transmitted by tick species such as Rhipicephalus (Boophilus) microplus, blood-sucking insects, and fomites (needles, clippers, and other blood contaminated equipment). During the acute phase of infection, animals may develop fever, anemia, jaundice, and hepatosplenomegaly. The aims of this study are to quantify the bacteremia by quantitative PCR in eight naïve calves experimentally infected by A. marginale [splenectomized (n = 4), and intact/non-splenectomized (n = 4)], and to correlate these findings with markers of oxidative stress on days 0, 8, 15, 21 and 23 post-infection. Complete blood counts (CBC) were performed in both groups. Lipid peroxidation was estimated by quantifying thiobarbituric acid reactive substances (TBARS); and non-enzymatic antioxidants were assessed by erythrocyte content of non-protein thiols (NPSH). There were no significant differences in complete blood counts (CBC) between the two groups. However, both groups had a slight decrease on packet cell volume (PCV), erythrocytes and hemoglobin concentration, as well as an increase in total leukocyte counts due to elevated lymphocytes when comparing pre and post-infection with A. marginale. Progressive increase on TBARS levels and concomitant decrease on NPSH content were observed in all animals, without significant differences between splenectomized and intact animals. A positive correlation between bacteremia and TBARS, and a negative correlation between bacteremia and NPSH were observed in both groups with higher correlation for NPSH in splenectomized animals. A negative correlation between TBARS and NPSH levels was observed in both groups indicating lipid peroxidation without a non-enzymatic antioxidant response. The results of experimental infection by A. marginale in cattle showed that bacteremia has an impact on lipid peroxidation regardless of the splenectomy.

Superinfection Exclusion of the Ruminant Pathogen Anaplasma marginale in Its Tick Vector Is Dependent on the Time between Exposures to the Strains.

UNLABELLED: The remarkable genetic diversity of vector-borne pathogens allows for the establishment of superinfection in the mammalian host. To have a long-term impact on population strain structure, the introduced strains must also be transmitted by a vector population that has been exposed to the existing primary strain. The sequential exposure of the vector to multiple strains frequently prevents establishment of the second strain, a phenomenon termed superinfection exclusion. As a consequence, superinfection exclusion may greatly limit genetic diversity in the host population, which is difficult to reconcile with the high degree of genetic diversity maintained among vector-borne pathogens. Using Anaplasma marginale, a tick-borne bacterial pathogen of ruminants, we hypothesized that superinfection exclusion is temporally dependent and that longer intervals between strain exposures allow successful acquisition and transmission of a superinfecting strain. To test this hypothesis, we sequentially exposed Dermacentor andersoni ticks to two readily tick-transmissible strains of A. marginale The tick feedings were either immediately sequential or 28 days apart. Ticks were allowed to transmission feed and were individually assessed to determine if they were infected with one or both strains. The second strain was excluded from the tick when the exposure interval was brief but not when it was prolonged. Midguts and salivary glands of individual ticks were superinfected and transmission of both strains occurred only when the exposure interval was prolonged. These findings indicate that superinfection exclusion is temporally dependent, which helps to account for the differences in pathogen strain structure in tropical compared to temperate regions. IMPORTANCE: Many vector-borne pathogens have marked genetic diversity, which influences pathogen traits such as transmissibility and virulence. The most successful strains are those that are preferentially transmitted by the vector. However, the factors that determine successful transmission of a particular strain are unknown. In the case of intracellular, bacterial, tick-borne pathogens, one potential factor is superinfection exclusion, in which colonization of ticks by the first strain of a pathogen it encounters prevents the transmission of a second strain. Using A. marginale, the most prevalent tick-borne pathogen of cattle worldwide, and its natural tick vector, we determined that superinfection exclusion occurs when the time between exposures to two strains is brief but not when it is prolonged. These findings suggest that superinfection exclusion may influence strain transmission in temperate regions, where tick activity is limited by season, but not in tropical regions, where ticks are active for long periods.

Influence of experimental Anaplasma marginale infection and splenectomy on NTPDase and 5'nucleotidase activities in platelets of cattle.

The objective of this paper was to evaluate NTPDase and 5'-nucleotidase activities in platelets of bovine with and without spleen and infected by Anaplasma marginale. Our results demonstrate that infection along with splenectomy is able of inducing a profile of cellular protection, which showed an increase in the degradation of the nucleotides ATP and ADP by NTPDase, in addition to AMP by 5'nucleotidase to form the nucleoside adenosine in platelets, i.e., the enzymatic activities of platelets were increased in splenectomized animals when compared to non-splenectomized group. It notes that adenosine is a molecule with anti-inflammatory function. But this profile is related to a deficiency in immune signaling triggered by nucleotide ATP, which may be related to the increase in bacteremia and disability in combating the parasite in splenectomized host.

Molecular and immunological characterization of three strains of Anaplasma marginale grown in cultured tick cells.

Anaplasma marginale is an economically important tick-borne pathogen of cattle that causes bovine anaplasmosis. A wide range of geographic strains of A. marginale have been isolated from cattle, several of which have been characterized using genomics and proteomics. While many of these strains have been propagated in tick lines, comparative analyses after propagation in tick cells have not been reported. The overall purpose of this research therefore was to compare the degree of conservation of selected genes after propagation in tick cell culture among A. marginale strains from the U.S. (the Virginia strain) and Brazil (UFMG1 and UFMG2 strains). The genes studied herein included those which encode the proteins HSP70 and SODB involved in heat shock and stress responses, respectively, and two genes that encode major surface proteins MSP4 and MSP5. Strain identities were first confirmed by sequencing the tandem repeats of the msp1a gene which encodes for the adhesin, MSP1a. The results of these studies demonstrated that the genes encoding for both stress response and heat shock proteins were highly conserved among the three A. marginale strains. Antibodies specific for MSP4, MSP5, SODB and HSP70 proteins were used to further characterize the A. marginale strains, and they reacted with all of these strains propagated in tick cell culture, providing further evidence for antigenic conservation. Although antigenic differences were not found among the three A. marginale strains, multi-locus sequence analysis (MLSA) performed with nucleotide sequences of these genes demonstrated that the A. marginale Brazilian and U.S. strains fall in different clades. These results showed that phylogenetically distant strains of A. marginale are antigenically conserved, even after several in vitro passages, supporting the use of some of the above conserved proteins as candidates for universal vaccines.

Reduced Infectivity in cattle for an outer membrane protein mutant of Anaplasma marginale.

Anaplasma marginale is the causative agent of anaplasmosis in cattle. Transposon mutagenesis of this pathogen using the Himar1 system resulted in the isolation of an omp10 operon insertional mutant referred to as the omp10::himar1 mutant. The work presented here evaluated if this mutant had morphological and/or growth rate defects compared to wild-type A. marginale. Results showed that the morphology, developmental cycle, and growth in tick and mammalian cell cultures are similar for the mutant and the wild type. Tick transmission experiments established that tick infection levels with the mutant were similar to those with wild-type A. marginale and that infected ticks successfully infected cattle. However, this mutant exhibited reduced infectivity and growth in cattle. The possibility of transforming A. marginale by transposon mutagenesis coupled with in vitro and in vivo assessment of altered phenotypes can aid in the identification of genes associated with virulence. The isolation of deliberately attenuated organisms that can be evaluated in their natural biological system is an important advance for the rational design of vaccines against this species.

Transcriptional pathways associated with the slow growth phenotype of transformed Anaplasma marginale.

BACKGROUND: The ability to genetically manipulate bacteria has been fundamentally important for both basic biological discovery and translational research to develop new vaccines and antibiotics. Experimental alteration of the genetic content of prokaryotic pathogens has revealed both expected functional relationships and unexpected phenotypic consequences. Slow growth phenotypes have been reported for multiple transformed bacterial species, including extracellular and intracellular pathogens. Understanding the genes and pathways responsible for the slow growth phenotype provides the opportunity to develop attenuated vaccines as well as bacteriostatic antibiotics. Transformed Anaplasma marginale, a rickettsial pathogen, exhibits slow growth in vitro and in vivo as compared to the parent wild type strain, providing the opportunity to identify the underlying genes and pathways associated with this phenotype. RESULTS: Whole genome transcriptional profiling allowed for identification of specific genes and pathways altered in transformed A. marginale. Genes found immediately upstream and downstream of the insertion site, including a four gene operon encoding key outer membrane proteins, were not differentially transcribed between wild type and transformed A. marginale. This lack of significant difference in transcription of flanking genes and the large size of the insert relative to the genome were consistent with a trans rather than a cis effect. Transcriptional profiling across the complete genome identified the most differentially transcribed genes, including an iron transporter, an RNA cleaving enzyme and several genes involved in translation. In order to confirm the trend seen in translation-related genes, K-means clustering and Gene Set Enrichment Analysis (GSEA) were applied. These algorithms allowed evaluation of the behavior of genes as groups that share transcriptional status or biological function. Clustering and GSEA confirmed the initial observations and found additional pathways altered in transformed A. marginale. Three pathways were significantly altered as compared to the wild type: translation, translation elongation, and purine biosynthesis. CONCLUSIONS: Identification of perturbed genes and networks through genome wide transcriptional profiling highlights the relevance of pathways such as nucleotide biosynthesis, translation, and translation elongation in the growth phenotype of obligate intracellular bacteria. These genes and pathways provide specific targets for development of slow growing attenuated vaccines and for bacteriostatic antibiotics.

In vitro cultivation of Anaplasma marginale and A. phagocytophilum in tick cell lines: a review.

Continuous cell lines have been established from several ixodid and argasid tick species, representing an excellent tool suitable for the isolation of pathogens and their subsequent propagation, which in turn allows the production of antigenic material for diagnostic tests, antibody and vaccine production, and also for studies on host-vector-pathogen relationships. This paper reviews the use of tick cells for culture initiation and maintenance of two obligate intracellular bacterial pathogens, Anaplasma marginale and Anaplasma phagocytophilum. These in vitro cultivation systems have been used in a wide range of studies, covering morphological ultrastructural analysis, genetics, proteomics and biological differences between strains, including genome transcriptional and protein expression approaches, enabling comparisons between host and vector cells. Thus, such systems open a new window for a better understanding of interactions between pathogens and tick cells. Last but not least, such systems contribute to the reduction in usage of animals for experimental research, as antigenic material can be produced in reasonably large quantities without the use of in vivo species-specific systems.

In vitro cultivation of Anaplasma marginale and A. phagocytophilum in tick cell lines: a review / Cultivo in vitro de Anaplasma marginale e A. phagocytophilum em células de carrapatos: uma revisão

Continuous cell lines have been established from several ixodid and argasid tick species, representing an excellent tool suitable for the isolation of pathogens and their subsequent propagation, which in turn allows the production of antigenic material for diagnostic tests, antibody and vaccine production, and also for studies on host-vector-pathogen relationships. This paper reviews the use of tick cells for culture initiation and maintenance of two obligate intracellular bacterial pathogens, Anaplasma marginale and Anaplasma phagocytophilum. These in vitro cultivation systems have been used in a wide range of studies, covering morphological ultrastructural analysis, genetics, proteomics and biological differences between strains, including genome transcriptional and protein expression approaches, enabling comparisons between host and vector cells. Thus, such systems open a new window for a better understanding of interactions between pathogens and tick cells. Last but not least, such systems contribute to the reduction in usage of animals for experimental research, as antigenic material can be produced in reasonably large quantities without the use of in vivo species-specific systems.

Linhagens contínuas de células já foram estabelecidas a partir de várias espécies de carrapatos ixodídeos e argasídeos e representam uma ferramenta excelente para o isolamento e propagação de patógenos, permitindo a produção de material antigênico para testes diagnósticos, produção de anticorpos e vacinas, e também para estudos das relações entre hospedeiro-vetor-patógenos. Este artigo revisa o uso de células de carrapatos para estabelecimento e manutenção in vitro de dois patógenos intracelulares, Anaplasma marginale e Anaplasma phagocytophilum. Estes sistemas de cultivo in vitro, têm sido utilizados em vários estudos, tais como análises morfológicas, genéticas, proteômicas e estudos diferenciais entre isolados, incluindo genômica transcricional e expressões proteicas, permitindo comparações entre células dos hospedeiros e dos vetores. Tais sistemas constituem, portanto, uma nova abordagem para melhor entendimento das relações entre patógenos e células de carrapatos. Além disso, tais sistemas contribuem para a redução do uso de animais de experimentação, uma vez que permitem a produção de grandes quantidades de material antigênico sem o uso de sistemas espécie-específicos in vivo.

Expression patterns of Anaplasma marginale Msp2 variants change in response to growth in cattle, and tick cells versus mammalian cells.

Antigenic variation of major surface proteins is considered an immune-evasive maneuver used by pathogens as divergent as bacteria and protozoa. Likewise, major surface protein 2 (Msp2) of the tick-borne pathogen, Anaplasma marginale, is thought to be involved in antigenic variation to evade the mammalian host immune response. However, this dynamic process also works in the tick vector in the absence of immune selection pressure. We examined Msp2 variants expressed during infection of four tick and two mammalian cell-lines to determine if the presence of certain variants correlated with specific host cell types. Anaplasma marginale colonies differed in their development and appearance in each of the cell lines (P<0.001). Using Western blots probed with two Msp2-monospecific and one Msp2-monoclonal antibodies, we detected expression of variants with differences in molecular weight. Immunofluorescence-assay revealed that specific antibodies bound from 25 to 60% of colonies, depending on the host cell-line (P<0.001). Molecular analysis of cloned variant-encoding genes demonstrated expression of different predominant variants in tick (V1) and mammalian (V2) cell-lines. Analysis of the putative secondary structure of the variants revealed a change in structure when A. marginale was transferred from one cell-type to another, suggesting that the expression of particular Msp2 variants depended on the cell-type (tick or mammalian) in which A. marginale developed. Similarly, analysis of the putative secondary structure of over 200 Msp2 variants from ticks, blood samples, and other mammalian cells available in GenBank showed the predominance of a specific structure during infection of a host type (tick versus blood sample), demonstrating that selection of a possible structure also occurred in vivo. The selection of a specific structure in surface proteins may indicate that Msp2 fulfils an important role in infection and adaptation to diverse host systems. Supplemental Abstract in Spanish (File S1) is provided.

Expression of Anaplasma marginale ankyrin repeat-containing proteins during infection of the mammalian host and tick vector.

Transmission of tick-borne pathogens requires transition between distinct host environments with infection and replication in host-specific cell types. Anaplasma marginale illustrates this transition: in the mammalian host, the bacterium infects and replicates in mature (nonnucleated) erythrocytes, while in the tick vector, replication occurs in nucleated epithelial cells. We hypothesized that proteins containing ankyrin motifs would be expressed by A. marginale only in tick cells and would traffic to the infected host cell nucleus. A. marginale encodes three proteins containing ankyrin motifs, an AnkA orthologue (the AM705 protein), AnkB (the AM926 protein), and AnkC (the AM638 protein). All three A. marginale Anks were confirmed to be expressed during intracellular infection: AnkA is expressed at significantly higher levels in erythrocytes, AnkB is expressed equally by both infected erythrocytes and tick cells, and AnkC is expressed exclusively in tick cells. There was no evidence of any of the Ank proteins trafficking to the nucleus. Thus, the hypothesis that ankyrin-containing motifs were predictive of cell type expression and nuclear localization was rejected. In contrast, AnkA orthologues in the closely related A. phagocytophilum and Ehrlichia chaffeensis have been shown to localize to the host cell nucleus. This difference, together with the lack of a nuclear localization signal in any of the AnkA orthologues, suggests that trafficking may be mediated by a separate transporter rather than by endogenous signals. Selection for divergence in Ank function among Anaplasma and Ehrlichia spp. is supported by both locus and allelic analyses of genes encoding orthologous proteins and their ankyrin motif compositions.

Silencing of genes involved in Anaplasma marginale-tick interactions affects the pathogen developmental cycle in Dermacentor variabilis.

BACKGROUND: The cattle pathogen, Anaplasma marginale, undergoes a developmental cycle in ticks that begins in gut cells. Transmission to cattle occurs from salivary glands during a second tick feeding. At each site of development two forms of A. marginale (reticulated and dense) occur within a parasitophorous vacuole in the host cell cytoplasm. However, the role of tick genes in pathogen development is unknown. Four genes, found in previous studies to be differentially expressed in Dermacentor variabilis ticks in response to infection with A. marginale, were silenced by RNA interference (RNAi) to determine the effect of silencing on the A. marginale developmental cycle. These four genes encoded for putative glutathione S-transferase (GST), salivary selenoprotein M (SelM), H+ transporting lysosomal vacuolar proton pump (vATPase) and subolesin. RESULTS: The impact of gene knockdown on A. marginale tick infections, both after acquiring infection and after a second transmission feeding, was determined and studied by light microscopy. Silencing of these genes had a different impact on A. marginale development in different tick tissues by affecting infection levels, the densities of colonies containing reticulated or dense forms and tissue morphology. Salivary gland infections were not seen in any of the gene-silenced ticks, raising the question of whether these ticks were able to transmit the pathogen. CONCLUSION: The results of this RNAi and light microscopic analyses of tick tissues infected with A. marginale after the silencing of genes functionally important for pathogen development suggest a role for these molecules during pathogen life cycle in ticks.

Quantitative differences in salivary pathogen load during tick transmission underlie strain-specific variation in transmission efficiency of Anaplasma marginale.

The relative fitness of arthropod-borne pathogens within the vector can be a major determinant of pathogen prevalence within the mammalian host population. Strains of the tick-borne rickettsia Anaplasma marginale differ markedly in transmission efficiency, with a consequent impact on pathogen strain structure. We have identified two A. marginale strains with significant differences in the transmission phenotype that is effected following infection of the salivary gland. We have proposed competing hypotheses to explain the phenotypes: (i) both strains are secreted equally, but there is an intrinsic difference in infectivity for the mammalian host, or (ii) one strain is secreted at a significantly higher level and thus represents delivery of a greater pathogen dose. Quantitative analysis of pathogen replication and secretion revealed that the high-efficiency St. Maries strain replicated to a 10-fold-higher titer and that a significantly greater percentage of infected ticks secreted A. marginale into the saliva and did so at a significantly higher level than for the low-efficiency Israel vaccine strain. Furthermore, the transmission phenotype of the vaccine strain could be restored to that of the St. Maries strain simply by increasing the delivered pathogen dose, either by direct inoculation of salivary gland organisms or by increasing the number of ticks during transmission feeding. We identified morphological differences in the colonization of each strain within the salivary glands and propose that these reflect strain-specific differences in replication and secretion pathways linked to the vector-pathogen interaction.

Genetic diversity of major surface protein 1a of Anaplasma marginale in beef cattle.

The present study was aimed to demonstrate genotypic diversity of Anaplama marginale in infected beef herds grazing within anaplasmosis endemic regions. The genotypic diversity was identified among different herds, within each herd, and also within single animals. The Israeli strains revealed unique characteristics of MSP1a repeats and, in addition to the published repeats, six new tandem repeats designated Is1-5, and Is9 were identified. The superinfections of individual Anaplama centrale vaccinated animals with two genotypically different A. marginale strains were detected. Six out of 43 vaccinated animals in the G herd were each infected with two A. marginale strains carrying two distinct genotypes; in this herd the follow-up during years 2003-2007 demonstrated that several animals carried different msp1a genotypes at different time points. Coinfection with two different genotypes of A. marginale in A. centrale vaccinated cattle was observed in another herd, as well. It appears that A. marginale is composed of a heterogeneous changing bacterial population that evolves in the host or, the genotypic diversity implies high transmission intensity by the vector, or both. Learning how this diversity is generated and identification of distinct A. marginale strains coupled with high sequence variation of MSP1a will aid in understanding Anaplasma transmission and disease development.

Studies on correlations among parasitaemia and some hemolytic indices in two tropical diseases (theileriosis and anaplasmosis) in Fars province of Iran.

This study was carried out in two observational clinical studies. Study 1 comprised 50 adult crossbred cattle naturally infected by Theileria annulata. Infected animals were divided into 4 subgroups with different parasitaemia (<1%, 1-3%, 3-5% and >5%). Study 2 comprised 20 adult crossbred cattle naturally infected by Anaplasma marginale. Infected animals were divided into 3 subgroups with different parasitaemia (<10%, 10-20% and 20-30%). In study 1, a significant negative correlation (P<0.001) was observed between parasitaemia and superoxide dismutase (SOD). Positive correlations (P<0.001) were observed between parasitaemia and lactate dehydrogenase (LDH) and mean corpuscular fragility (MCF). In study 2 positive correlations (P<0.05) were observed among parasitaemia and MCF and LDH activity. SOD activity had a negative correlation with parasitaemia in cattle with parasitaemia lower than 10% but no significant correlation (P>0.05) was observed between SOD activity and parasitaemia in cattle with 10-20 and 20-30% parasitaemia. In comparison of both studies we came to the conclusion that in theileriosis as the severity of disease increased the anaemia, MCF and LDH activity increased and SOD activity decreased at any parasitaemia, but in anaplasmosis the anaemia, MCF and LDH activity increased at any parasitaemia but SOD activity decreased only in early but not in advanced stages of disease.

Experimental transmission of Anaplasma marginale by male Dermacentor reticulatus.

BACKGROUND: Bovine anaplasmosis has been reported in several European countries, but the vector competency of tick species for Anaplasma marginale from these localities has not been determined. Because of the wide distributional range of Dermacentor reticulatus within Europe and the major role of Dermacentor spp. as a vector of A. marginale in the United States, we tested the vector competency of D. reticulatus for A. marginale. RESULTS: Male D. reticulatus were allowed to feed for 7 days on a calf persistently infected with a Zaria isolate of A. marginale, after which they were removed and held off-host for 7 days. The ticks were then allowed to feed a second time for 7 days on a susceptible tick-naïve calf. Infection of calf No. 4291 was detected 20 days post exposure (p.i.) and confirmed by msp4 PCR. Thirty percent of the dissected acquisition fed ticks was infected. In addition, A. marginale colonies were detected by light microscopy in the salivary glands of the acquisition fed ticks. Transmission of A. marginale to calf No. 9191 was confirmed by examination of Giemsa-stained blood smears and msp4 PCR. Ticks were dissected after transmission feeding and presence of A. marginale was confirmed in 18.5% of the dissected ticks. CONCLUSION: This study demonstrates that D. reticulatus males are competent vectors of A. marginale. Further studies are needed to confirm the vector competency of D. reticulatus for other A. marginale strains from geographic areas in Europe.

In vivo endothelial cell infection by Anaplasma marginale.

Anaplasma marginale has recently been shown to infect endothelial cells in vitro, but it remains unknown as to whether endothelial infection also occurs in vivo. In this report, we demonstrate through dual fluorescence microscopy that A marginale, detected by the monoclonal antibody ANAF16C1, co-localizes with the endothelial cell marker, von Willebrand factor, in tissue sections from an experimentally inoculated calf. The results indicate that A marginale infection includes endothelial cells and has implications for both pathogenesis and immune mechanisms.

Efficacy of enrofloxacin against severe experimental Anaplasma marginale infections in splenectomized calves.

Four Anaplasma marginale-infected splenectomized calves with greater than 25% parasitized erythrocytes received enrofloxacin at 12.5 mg/kg SC twice, 48 hours apart. Two infected splenectomized calves were designated as untreated controls. A precipitous decline in percent parasitized erythrocytes from 39.13% to less than 1% was observed over 12 days following treatment. However, a self-limiting recrudescence of A. marginale parasites was observed within 30 days after treatment. Untreated control calves became moribund and were euthanized. These data indicate that the regimen of enrofloxacin tested herein ameliorates, but does not eliminate, A. marginale infections in splenectomized calves.

A longitudinal study of sero-conversion to tick-borne pathogens in smallholder dairy youngstock in Tanzania.

A longitudinal study of sero-conversion of youngstock to the tick-borne pathogens Theileria parva, T. mutans, Anaplasma marginale, Babesia bigemina and B. bovis was conducted over two years on smallholder dairy farms in Tanga region, Tanzania. There was evidence of maternal antibodies to all tick-borne pathogens in animals less than 18 weeks of age. Seroprevalence increased as expected with age in animals older than this but seroprevalence profiles underestimated the force of infection due to waning antibody levels between samplings. By the end of the 2-year study, less than 50% of study animals had seroconverted to each of the tick-borne pathogens investigated, consistent with the low levels of tick attachment observed on the study animals. Some associations between seroconversion to tick-borne pathogens, and counts of their known tick vectors on the animals, were identified as expected. However, some were not, suggesting that counts of some tick species may act as an index of rates of attachment of other vector species. Variation in acaricide treatment frequencies was not associated with variations in tick-borne pathogen seroprevalence suggesting that acaricides may be used more frequently than necessary on many farms. Most animals were zero-grazed, a management system associated with a significantly lower likelihood that animals seroconverted to any tick-borne pathogen except A. marginale. Seroprevalence varied locally with farm location (particularly for Babesia spp.) but was not well predicted by indices of ecological conditions. Our findings suggest that attempts to achieve a state of 'endemic stability' for tick-borne pathogens may be unreasonable on the smallholder dairy farms studied but reductions in the frequency of use of acaricides may be possible following prospective studies of effects on mortality and morbidity due to tick-borne pathogens.

The impact of 2 dipping systems on endemic stability to bovine babesiosis and anaplasmosis in cattle in 4 communally grazed areas in Limpopo Province, South Africa.

A 12-month study was conducted in 4 communal grazing areas in the Bushbuckridge region, Limpopo Province, South Africa. The main objective was to investigate the impact of reduced acaricide application on endemic stability to bovine babesiosis (Babesia bigemina and Babesia bovis) and anaplasmosis (Anaplasma marginale) in the local cattle population. To this end 60 cattle in each communal grazing area were bled at the beginning and the conclusion of the experimental period and their sera were assayed for B. bovis, B. bigemina and Anaplasma antibodies. Cattle in the intensively dipped group were dipped 26 times and maintained on a 14-day dipping interval throughout the study, whereas cattle in the strategically dipped group were dipped only 13 times. Three cattle, from which adult ticks were collected, were selected from each village, while immature ticks were collected by drag-sampling the surrounding vegetation. During the dipping process, a questionnaire aimed at assessing the prevalence of clinical cases of tick-borne disease, abscesses and mortalities was completed by an Animal Health Technician at each diptank. An increase in seroprevalence to B. bovis and B. bigemina and a decrease in seroprevalence to Anaplasma was detected in the strategically dipped group while in the intensively dipped group the converse was true. Amblyomma hebraeum was the most numerous tick species on the cattle, and Rhipicephalus (Boophilus) microplus was more plentiful than Rhipicephalus (Boophilus) decoloratus. Drag samples yielded more immature stages of A. hebraeum than of Rhipicephalus (Boophilus) spp. The incidence of clinical cases of tick-borne disease and of abscesses increased in the strategically dipped group at the start of the survey.