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PubMed1993

A comparison of field tick infestation on N'Dama, Zebu and N'Dama x Zebu crossbred cattle.

Tick burdens, estimated from cumulative tick counts, were determined on N'Dama, Zebu and F1 N'Dama x Zebu crossbred cattle. N'Dama showed significantly fewer ticks than the Zebu and F1 cattle (P < 0.001). A previous trypanosomosis infection did not affect tick burdens on the three genotypes. Amblyomma variegatum had a prevalence of 84.8% and Hyalomma spp. 15.2%, with the ratio similar on all cattle breeds. Trypanosomosis infection increased the serological prevalence of Anaplasma marginale in Zebu and F1 cattle but not in N'Dama cattle. These observations supported the view that N'Dama cattle are less susceptible to tick attachment than Zebu cattle.

Animals↗

PubMed1995

Review of the molecular biology of Cowdria ruminantium.

Cowdria ruminantium is a rickettsial agent which causes heartwater, an economically important disease of livestock in the tropics and the Caribbean. Significant advances have been made in the molecular biology of C. ruminantium since its reproducible propagation in vitro in bovine endothelial cells. These advances have been targeted towards the development of improved vaccines and diagnostic tests. Several immunogenic proteins of C. ruminantium have been identified and monoclonal antibodies have been developed to some. The gene for the 21 kDa C. ruminantium protein has been cloned, characterized, sequenced and expressed to high levels to produce a recombinant analogue. This gene is conserved amongst all C. ruminantium isolates tested. The gene for the immunodominant 32 kDa protein has also been cloned recently. Analysis shows that this protein varies structurally between different C. ruminantium isolates. Recombinant protein analogues will have application in vaccine studies and subunit enzyme-linked immunosorbent assay (ELISA) developments. In the field of serology, a 32 kDa protein specific competitive ELISA (cELISA) is available and a 21 kDa protein specific direct and a cELISA is being developed. Detection of C. ruminantium in Amblyomma ticks and in animals is now possible using DNA and RNA probes and the sensitivity of these nucleic acid based assays is being maximized using polymerase chain reaction (PCR). Sequencing of the 16s rRNA gene has revealed the close phylogenetic relationship of C. ruminantium to Rickettsia, Anaplasma and Ehrlichia species. The implications of these studies will be discussed.

Animals↗

PubMed1995

Epidemiology of babesiosis and anaplasmosis in South and Central America.

Babesiosis (Babesia bovis and Babesia bigemina) and anaplasmosis (Anaplasma marginale) affect native cattle from Uruguay and northern Argentina to Guatemala. The distribution of Babesia follows the dissemination of the tick vector. Seroepidemiological studies showed that enzootic instability exist in herds in several areas of the region. However, clinical cases occurred less frequently than expected. The babesial tick infection rate is related to the vector abundance which in turn is regulated by climate. Bos indicus cattle, because of tick resistance, are less likely to be infected by the vector. This can result in herd instability, but clinically is partly compensated by the resistance of Bos indicus and their crosses to babesiosis. Excessive use of acaricides and rotational grazing appears to be related to outbreaks of babesiosis especially in dairy cattle. Factors involved in the epidemiology and transmission of anaplasmosis are not well defined. The role of ticks, haematophagous diptera, iatrogenic and intrauterine transmissions needs to be investigated under local conditions. Therefore, the knowledge of this disease is meager. Utilization of biotechnological methods may help in obtaining information on Babesia-Boophilus-Bos relationship and on the transmission of A. marginale.

Anaplasmosis↗

PubMed1995

Strategies for the control of one-host ticks and relationship with tick-borne diseases in South America.

A variety of hemoparasites occurs in ruminants of South America (12 degrees 00'N-56 degrees 00'S), but there is consensus on the significant economic impact of babesiosis (Babesia bovis and Babesia bigemina) and anaplasmosis (Anaplasma marginale). The tick vector for the Babesia spp. for the whole region is Boophilus microplus which has been the most important target in control/eradication programs. Boophilus microplus and many different hematophagous diptera species are considered vectors of A. marginale. In areas where babesiosis and anaplasmosis are present, the wide range of climates (tropical, subtropical, temperate), husbandry practices (subsistence, extensive, intensive), and tick control strategies has led to the concepts of enzootic stability and instability. Within this framework, six control strategies for Boophilus microplus can be identified: (1) absolute tick control--with the establishment of quarantine areas and the implementation of state legislation; (2) absolute tick control in marginal areas--using local epidemiological knowledge, strategic treatments, quarantine areas and the intervention of the state; (3) prophylactic treatment--with a predetermined acaricide treatment schedule aimed at reducing tick burdens to an acceptably low level; (4) threshold treatment--acaricides are applied by the farmer whenever the number of ticks on cattle exceed a threshold level; (5) opportunistic treatment--acaricides are applied when cattle are assembled for other purposes; (6) integrated tick control--principally involving the combination of acaricides with the use of tick-resistant crossbred cattle.

Anaplasmosis↗

PubMed1995

Vaccine development from a commercial point of view.

The development of a commercial vaccine comprises distinct stages. Initiation of a research project is triggered by demands from the market. If commercial and technical requirements are met, a feasibility study is carried out. Research is started, and aimed at formulating the product profile (what the product looks like). The product profile is subject to requirements set by the market (e.g. whether the product will fit into existing vaccination schedules) and very often technical aspects affect the product profile (e.g. whether the freeze-dried product is easy to reconstitute). Only after a cost-profit analysis is positive, the development phase is entered. During this phase, experiments are carried out to obtain registration. After the product has been registered it is ready for production and marketing. Only few vaccines for hemoparasitic diseases have reached the market. These comprise: attenuated parasites (Toxoplasma gondii, Eimeria species); killed vaccines (Anaplasma marginale) and subunit vaccines (Babesia canis). Factors relating to the product potential of these vaccines are discussed.

Animals↗

PubMed1996

Piroplasms of domestic animals in the Macedonia region of Greece. 3. Piroplasms of small ruminants.

A study was carried out on piroplasms of small ruminants in the Macedonia region of Greece. During 1984-1985, 721 serum samples were collected from sheep in 49 localities and 487 from goats in 43 localities. Blood smears were also prepared from 26 sheep and eight goats in 16 localities. The prevalence of positive IFA titres for sheep and goat sera, respectively, was: 24.6% and 0.6% for Theileria ovis, 52.1% and 36.4% for Babesia ovis, 10.5% and 4.2% for Babesia motasi, 12.6% and 6.6% for Babesia crassa. Many of the positive reactions, specially for the large Babesia species, B. motasi and B. crassa, probably resulted from cross-reactions and the presence of these two species is doubtful. Antibodies against B. ovis were found in animals from more than 90% of the localities. T. ovis was also common, but it was confined essentially to sheep. The examination of Giemsa stained blood smears revealed the presence of T. ovis and B. ovis in sheep, and Anaplasma ovis in a goat.

Animals↗

PubMed1991

Competitive enzyme-linked immunosorbent assay for heartwater using monoclonal antibodies to a Cowdria ruminantium-specific 32-kilodalton protein.

Hybridomas producing monoclonal antibodies (mAb) to Cowdria ruminantium were raised. Four mAbs of the IgG isotype reacted in western blots with a 32-kilodalton Cowdria protein (Cr32), which had previously been shown to be conserved and immunodominant. A fifth mAb of the IgM isotype recognized a 40-kDa Cowdria protein. The latter mAb was negative in an indirect fluorescent antibody test (IFA), whereas the other four were positive. mAb No. 4F10B4 showed the strongest signal in western blots using three different stocks of Cowdria. Immuno-gold labeling of Cowdria organisms in vitro using 4F10B4 showed that Cr32 has surface-exposed antigenic determinants. Using mAb 4F10B4, a competitive ELISA was developed which detected specific Cowdria antibodies in goat, sheep and cattle sera. Antibodies in animal sera competed with binding of mAb 4F10B4 to a crude sonicated Cowdria antigen obtained from infected endothelial cell cultures. The competition ELISA (CELISA) detected antibodies in 55 out of 70 (79%) goats experimentally infected with one of eight different Cowdria stocks. Fourteen out of the 15 sera which were shown negative in the CELISA were also negative in the IFA. Nevertheless, all 15 sera recognized some epitopes of the immunodominant Cowdria-specific 32 kDa protein as judged from their reaction with this protein in western blots. Overall, there was 89% agreement between CELISA and IFA considering all 70 goat sera. Moreover, antibodies were detected in nine out of nine sheep infected with one of three different stocks of Cowdria and in sera from calves experimentally infected by two different strains of heartwater. There were no cross-reactions with Ehrlichia phagocytophila antibodies in goat sera, nor with Anaplasma marginale antibodies in bovine sera. Lack of cross-reactivity and detection of antibodies to eight geographically widely distributed stocks of Cowdria, makes the competition ELISA a promising test for use in heartwater endemic areas.

Animals↗

PubMed1995

Phylogenetic analysis of the genus Rickettsia by 16S rDNA sequencing.

Rickettsiae are Gram-negative bacteria which multiply only inside host cells and need arthropods either as reservoirs or as vectors. Using the polymerase chain reaction and an automated laser fluorescent DNA sequencer, we amplified and sequenced the 16S rRNA (rDNA) of all available bacteria of the genus Rickettsia. R. tsutsugamushi remained close to the other bacteria of the genus Rickettsia using this technique, contrary to previous conclusions based on the study of the Sta-58 protein antigen. We found that R. canada was not included in the typhus group, as is currently recognized, but was grouped with the rickettsiae of the spotted fever group (SFG). All the SFG rickettsiae tested were grouped in the same cluster (R. conorii, Indian tick typhus rickettsia, Astrakhan fever rickettsia, Israeli tick typhus rickettsia, HA-91, R. sibirica, R. parkeri, "R. africae", "R. slovaca", R. rickettsii, Thai tick typhus rickettsia, R. japonica, R. massiliae, R. rhipicephali, R. montana, two recent isolates GS and Bar 29, R. australis, R. akari, R. bellii and R. helvetica). The recently described ELB bacterium, the agent of the Californian murine typhus, and AB bacterium, a bacterium associated with male killing in the ladybird beetle, were found in this cluster. The sequences of R. conorii Moroccan strain/Indian tick typhus rickettsia, R. massiliae/GS and R. sibirica/HA-91 were identical. All the rickettsiae had a unique ancestor with bacteria also isolated in arthropods (Ehrlichia, Cowdria, Anaplasma, Wolbachia pipientis), eventually pathogenic for mammals and implicated in parthenogenesis and cytoplasmic incompatibility. We conclude that a unique bacterium started a stable association with arthropod ancestors and generated the observed diversity of the currently isolated members of the Rickettsiales.

Base Sequence↗

PubMed2006

Bmcystatin, a cysteine proteinase inhibitor characterized from the tick Boophilus microplus.

The bovine tick Rhipicephalus (Boophilus) microplus is a blood-sucking animal, which is responsible for Babesia spp and Anaplasma marginale transmission for cattle. From a B. microplus fat body cDNA library, 465 selected clones were sequenced randomly and resulted in 60 Contigs. An open reading frame (ORF) contains 98 amino acids named Bmcystatin, due to 70% amino acid identity to a classical type 1 cystatin from Ixodes scapularis tick (GenBank Accession No. ). The Bmcystatin amino acid sequence analysis showed two cysteine residues, theoretical pI of 5.92 and M(r) of 11 kDa. Bmcystatin gene was cloned in pET 26b vector and the protein expressed using bacteria Escherichia coli BL21 SI. Recombinant Bmcystatin (rBmcystatin) purified by affinity chromatography on Ni-NTA-agarose column and ionic exchange chromatography on HiTrap Q column presented molecular mass of 11 kDa, by SDS-PAGE and the N-terminal amino acid sequenced revealed unprocessed N-terminal containing part of pelB signal sequence. Purified rBmcystatin showed to be a C1 cysteine peptidase inhibitor with K(i) value of 0.1 and 0.6 nM for human cathepsin L and VTDCE (vitellin degrading cysteine endopeptidase), respectively. The rBmcystatin expression analyzed by semi-quantitative RT-PCR confirmed the amplification of a specific DNA sequence (294 bp) in the fat body and ovary cDNA preparation. On the other hand, a protein band was detected in the fat body, ovary, and the salivary gland extracts using anti-Bmcystatin antibody by Western blot. The present results suggest a possible role of Bmcystatin in the ovary, even though the gene was cloned from the fat body, which could be another site of this protein synthesis.

Amino Acid Sequence↗

PubMed2004

Emerging intracellular bacterial infections.

Human infections by intracellular bacteria have been recognized for many years, but much of what we know about the pathogenesis of these diseases and their etiologic organisms has emerged within the past few years as a result of improved molecular-based means for their detection and classification. New insights concerning the epidemiology and pathogenesis of intracellular bacterial infections and methods for the detection of Chlamydophila pneumoniae, Ehrlichia chaffeensis, Anaplasma phagocytophilum, and Rickettsia species have made an impact on how we view them as agents of human disease. Emerging evidence suggesting a possible intracellular existence for another organism, Mycoplasma pneumoniae, may explain how this organism interacts with the host to induce chronic inflammatory conditions of the respiratory tract.

Chlamydia Infections↗

PubMed2006

Transcriptome analysis of the salivary glands of Dermacentor andersoni Stiles (Acari: Ixodidae).

Amongst blood-feeding arthropods, ticks of the family Ixodidae (hard ticks) are vectors and reservoirs of a greater variety of infectious agents than any other ectoparasite. Salivary glands of ixodid ticks secrete a large number of pharmacologically active molecules that not only facilitate feeding but also promote establishment of infectious agents. Genomic, proteomic and immunologic characterization of bioactive salivary gland molecules are, therefore, important as they offer new insights into molecular events occurring at the tick-host interface and they have implications for development of novel control strategies. The present work uses complementary DNA (cDNA) sequence analysis to identify salivary gland transcripts expressed by the Rocky Mountain wood tick, Dermacentor andersoni, a vector of the human pathogens causing Rocky Mountain spotted fever, Colorado tick fever, tularemia, and Powassan encephalitis as well as the veterinary pathogen Anaplasma marginale. Dermacentor andersoni is also capable of inducing tick paralysis. Automated single-pass DNA sequencing was conducted on 1440 randomly selected cDNA clones from the salivary glands of adult female D. andersoni collected during the early stages of feeding (18-24h). Analysis of the expressed sequence tags (ESTs) resulted in 544 singletons and 218 clusters with more than one quality read and attempts were made to assign putative functions to tick genes based on amino acid identity to published protein databases. Approximately 25.6% (195) of the sequences showed limited or no homology to previously identified gene products. A number of novel sequences were identified which presented significant sequence similarity to mammalian genes normally associated with extracellular matrix (ECM), regulation of immune responses, tumor suppression, and wound healing. Several coding sequences possessed various degrees of homology to previously described proteins from other tick species. Preliminary nucleotide variation analysis of these and other tick sequences suggests extensive nucleotide diversity, which has implications for evolution of tick feeding. Intra-species diversity studies can be a promising tool for identifying sequence variations potentially associated with phenotypic traits affecting vector-host-pathogen interactions.

Amino Acid Sequence↗

PubMed2004

Application of a reverse line blot assay to the study of haemoparasites in cattle in Uganda.

Recent advances in genomic technology have focused many veterinary researchers on the possibility of producing one multivalent recombinant vaccine against all the haemoparasites that infect cattle in the tropics. Before such a vaccine is developed it is essential to define target cattle populations as well as the range of anti-pathogen vaccines required in order to control disease. To further this objective, we have evaluated a reverse line blot (RLB) assay, which simultaneously detects the principal tick transmitted protozoan and rickettsial cattle pathogens, in different epidemiological scenarios in Uganda. A critical question is the sensitivity, particularly in relation to detecting carrier animals. As Theileria parva is considered to be the most important pathogen in the region, we assessed the sensitivity of the RLB assay for T. parva and showed that 1-2 x 10(3) parasites per ml of blood could be detected-a level comparable with previously developed PCR methods and well below conventional microscopic detection. We applied the RLB assay to evaluate the differences in pathogen profiles between crossbred and indigenous cattle and show that there were different profiles, with a low prevalence of T. parva and Theileria taurotragi in the indigenous cattle compared to a high prevalence in the crossbred cattle. In contrast, we show higher prevalences of Theileria mutans and Theileria velifera in the indigenous compared to the crossbred cattle. Interestingly Anaplasma marginale, Babesia bovis and Babesia bigemina were of low prevalence but a high prevalence of Ehrlichia bovis was seen, raising the question of whether this rickettsial species could be pathogenic in cattle. Analysis of animals with clinical symptoms of East Coast Fever showed that, while T. parva is a major cause of these symptoms, T. mutans and possibly T. taurotragi and T. velifera, may also cause clinical disease. Overall, the results presented here highlight the complexity of tick-borne pathogen infections in cattle in Uganda.

Anaplasmosis↗

PubMed2004

How do neutrophils and pathogens interact?

Many pathogens can manipulate macrophages after phagocytosis yet are efficiently killed by neutrophils. This poses the question of whether neutrophils have mechanisms that enable them to specifically recognise pathogens and have pathogens evolved mechanisms to modulate neutrophil function? Here, we review recent work on neutrophils and their interaction with four different bacteria: Staphylococcus aureus, Helicobacter pylori, Anaplasma phagocytophilum and members of the Enterobacteriae family.

Animals↗

PubMed2004

Transfusion-transmitted tick-borne infections: a cornucopia of threats.

Over the past several decades, the frequency of contact between humans and ticks has increased dramatically. Concomitantly, several newly recognized tick-borne pathogens have emerged joining those already known to be transmitted by ticks. Together these factors have led to an enhanced public health awareness of ticks, tick-borne agents, and their associated diseases. Reports that several of these agents are transmitted by blood transfusion have raised concerns about blood safety. The primary agents of interest are members of the genus Babesia, but Anaplasma phagocytophilum, Rickettsia rickettsii, Colorado tick fever virus, and tick-borne encephalitis virus also have been transmitted by transfusion. In many cases, these agents and their diseases share common features including vectors, symptoms, and diagnosis. Unfortunately, they also share the common problem of insufficient epidemiologic and transmissibility data necessary for making informed decisions regarding potential blood safety interventions. Although further surveillance and epidemiologic studies of tick-borne agents are clearly needed, at present only the Babesia warrant consideration for active intervention; because donor management strategies based on risk-factor questions are inadequate, leukoreduction not effective for agents found in red cells and pathogen inactivation remains problematic for red cell products. Despite the present unavailability of screening assays, some form of serologic and nucleic acid testing may be justified for the Babesia. Given that interactions between humans and ticks are likely to increase in the future, vigilance is required as new and extant tick-borne agents pose potential threats to transfusion safety.

Humans↗

PubMed2005

Evaluation of the recombinant LipL32 in enzyme-linked immunosorbent assay for the serodiagnosis of bovine leptospirosis.

The recombinant leptospiral protein LipL32 was evaluated for use in the diagnosis of bovine leptospirosis by enzyme-linked immunosorbent assay (rLipL32 IgG ELISA). The microscopic agglutination test (MAT) of 150 serum samples from cattle suspected of leptospirosis showed that 125 (83.3%) samples had positive reciprocal agglutination titres, which ranged from 100 to 1600. The highest titres were observed for the serovars Hardjoprajitno and Bratislava. In the rLipL32 IgG ELISA, 83.3% of the samples were positive. The sensitivity of IgG ELISA for 125 bovine sera, which had MAT titres of greater than or equal to 100, was 100%. ELISA showed a specificity of 100% with 58 bovine sera, which were negative at a 1:50 dilution in the MAT for Leptospira interrogans serovars. When analytical specificity of the IgG ELISA was evaluted using 60 bovine serum samples from animals showing serum antibodies to other pathogens that cause abortion in cattle, such as Babesia sp., Anaplasma sp. and Brucella sp. and no cross-reaction was observed. The recombinant LipL32 IgG ELISA can be an alternative to the MAT for diagnosis of leptospiral infection in cattle.

Agglutination Tests↗

PubMed2002

Molecular diagnosis of theileriosis and heartwater in bovines in Africa.

The advent of the polymerase chain reaction (PCR) coupled with the specificity of deoxyribocucleic acid (DNA)-DNA hybridization has led to the development of specific and sensitive molecular diagnostic tests to detect and characterize the organisms that cause theileriosis and heartwater. Theileriosis is a widespread disease of wild and domestic ruminants caused by apicomplexan parasites of the genus Theileria. Species-specific variations in small subunit ribosomal ribonucleic acid genes (SSUrRNA) have been used to develop probes that can distinguish between Theileria species such as T. parva, T. annulata, T. mutans, T. buffeli and T. taurotragi. Routine application of this test has led to the discovery of previously unknown species, such as Theileria sp. (buffalo) which is apparently apathogenic to both buffalo and cattle, and Theileria sp. (sable) which is pathogenic to sable and possibly also to roan antelope. In addition, characterization probes located in the internal transcribed spacer (ITS) can be used to distinguish between most isolates of the causative agents of East Coast fever (T. p. parva) and Corridor disease (T. p. lawrencei). Heartwater is an economically important disease of livestock and some wild ruminants, caused by the intracellular rickettsial parasite Ehrlichia (ex Cowdria) ruminantium. DNA probes used to detect and characterize E. ruminantium isolates include SSUrRNA (16S) probes, the pCS20 probe and map1 probes. A panel of eight 16S probes has been developed for the detection of E. ruminantium and related Ehrlichia species. There are probes for 5 different E. ruminantium genotypes, one which will detect all 5 of these genotypes, one to detect any Ehrlichia species other than E. ruminantium, and one for any Anaplasma species. The pCS20 probe is specific for E. ruminantium and is the most sensitive of the probes for E. ruminantium detection, but it is not able to distinguish among the different genotypes. The map1 gene has also been used for diagnosis, but the extensive polymorphism of this gene means that it is most useful for characterization of different genotypes of the parasite. Routine application of these tests has led to the discovery of new genotypes that are probably not E. ruminantium but are probably new species of Ehrlichia.

Animals↗

PubMed1999

Vaccine strategies for Cowdria ruminantium infections and their application to other ehrlichial infections.

Suman Mahan and co-authors review the strategies applied to develop improved vaccines for Cowdria ruminantium infections (heartwater). Inactivated vaccines using cell-cultured C. ruminantium organisms combined with an adjuvant are capable of protecting goats, sheep and cattle against lethal C. ruminantium challenge. Immune responses induced with this vaccine, or after recovery from infection, target outer membrane proteins of C. ruminantium, in particular the major antigenic protein 1 (MAP-1). Genetic immunizations with the gene encoding MAP-1 induce protective T helper cell type 1 responses against lethal challenge in a mouse model. Similarly, homologues of MAP-1 in other phylogenetically and antigenically related ehrlichial agents such as Anaplasma marginale and Ehrlichia chaffeensis are also targets of protective responses. Given the antigenic similarities between the related ehrlichial agents, common strategies of vaccine development could be applied against these agents that cause infections of importance in animals and humans.

Anaplasmataceae↗

PubMed2000

Risk of babesiosis and anaplasmosis in different ecological zones of Santa Cruz Department, Bolivia.

A cross-sectional study was done of seroprevalence of Babesia bigemina, B.bovis, and Anaplasma marginale in cattle from eastern Bolivia, to characterise the risk of tick-borne disease in three ecological zones. Nineteen farms were sampled in the subtropical humid zone, 13 in the dry subtropical zone and nine in the lower western valleys of the Andean massif. The enzyme-linked immunosorbent assay was used. All three pathogens were widespread. For B. bovis, seroprevalences were high (75-78%) in the two subtropical zones which thus had low risk of disease from this infection; but the western valleys were endemically unstable with higher risk. For B. bigemina, seroprevalences were lower (24-57%) in the two subtropical zones and thus these areas were endemically unstable for disease from this infection. However, the seroprevalence of B. bigemina in the western valleys was too low (13%) for risk of disease in susceptible cattle to be considered high. For A. marginale, the seroprevalences in the two subtropical zones were low (19-32%) and very low (6%) in the western valleys suggesting all these zones were endemically unstable for anaplasmosis. Data for individual farms were analysed for risk of both forms of babesiosis; this showed low risk of disease in the subtropical humid zone, higher risk in the dry subtropical zone and variable risk in the western valleys.

Agriculture↗