In vitro isolation of Cowdria ruminantium from plasma of infected ruminants

Byrom, B.; Yunker, Conrad E.; Donovan, Paula; Smith, Gillian

doi:10.1016/0378-1135(91)90019-c

Cited by 29 publications

(16 citation statements)

References 4 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…However, in situations where proof of infection requires direct demonstration of C. ruminantium infection or isolation of the agent, xenodiagnosis should be used. This should preferably be done in small ruminants instead of mice due to easier ante-and postmortem confirmation of infection and the availability of simple techniques for isolation of the organism in cell culture from ruminant plasma (8). In addition, certain C. ruminantium isolates appear to be noninfective for mice (18), though other conclusive experiments on this aspect are required.…”

Section: Discussionmentioning

confidence: 99%

“…Confirmation of infection was obtained by examination of brain smears prepared from biopsies performed during the febrile reaction or postmortem (53,56). During the febrile stage of infection, plasma was collected from some of the animals and inoculated into bovine endothelial cell cultures to attempt isolation of C. ruminantium (8). Infection of the cultures was confirmed by examination of cell smears prepared after the development of cytopathic effects in the cell monolayers.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Detection of the Agent of Heartwater, Cowdria ruminantium , in Amblyomma Ticks by PCR: Validation and Application of the Assay to Field Ticks

Peter¹,

Barbet

Alleman

et al. 2000

J Clin Microbiol

View full text Add to dashboard Cite

We have previously reported that the pCS20 PCR detection assay forCowdria ruminantium, the causative agent of heartwater disease of ruminants, is more sensitive than xenodiagnosis and the pCS20 DNA probe for the detection of infection in the vectorAmblyomma ticks. Here, we further assessed the reliability of the PCR assay and applied it to field ticks. The assay detected DNA of 37 isolates of C. ruminantium originating from sites throughout the distribution of heartwater and had a specificity of 98% when infected ticks were processed concurrently with uninfected ticks. The assay did not detect DNA of Ehrlichia chaffeensis, which is closely related to C. ruminantium. PCR sensitivity varied with tick infection intensity and was high (97 to 88%) with ticks bearing 107 to 104 organisms but dropped to 61 and 28%, respectively, with ticks bearing 103 and 102 organisms. The assay also detected C. ruminantium in collections of Amblyomma hebraeum andAmblyomma variegatum field ticks from 17 heartwater-endemic sites in four southern African countries. Attempts at tick transmission of infection to small ruminants failed with four of these collections. The pCS20 PCR assay is presently the most characterized and reliable test for C. ruminantium in ticks and thus is highly useful for field and laboratory epidemiological investigations of heartwater.

show abstract

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Detection of the Agent of Heartwater, Cowdria ruminantium , in Amblyomma Ticks by PCR: Validation and Application of the Assay to Field Ticks

Peter¹,

Barbet

Alleman

et al. 2000

J Clin Microbiol

View full text Add to dashboard Cite

show abstract

“…ruminantium strains and animal infections. E. ruminantium organisms were cultivated in vitro in bovine endothelial cells, as described previously (8). Strains representing different geographic origins in Africa and the Caribbean were selected.…”

Section: Methodsmentioning

confidence: 99%

Diversity of Ehrlichia ruminantium Major Antigenic Protein 1-2 in Field Isolates and Infected Sheep

2009

View full text Add to dashboard Cite

Proteins expressed from the map1 multigene family of Ehrlichia ruminantium are strongly recognized by immune T and B cells from infected animals or from animals that were infected and have recovered from heartwater disease (although still remaining infected carriers). Analogous multigene clusters also encode the immunodominant outer membrane proteins (OMPs) in other ehrlichial species. Recombinant protein analogs of the expressed genes and DNA vaccines based on the multigene clusters have been shown to induce protective immunity, although this was less effective in heterologous challenge situations, where the challenge strain major antigenic protein 1 (MAP1) sequence differed from the vaccine strain MAP1. Recent data for several ehrlichial species show differential expression of the OMPs in mammalian versus tick cell cultures and dominant expression of individual family members in each type of culture system. However, many genes in the clusters appear to be complete and functional and to generate mRNA transcripts. Recent data also suggest that there may be a low level of protein expression from many members of the multigene family, despite primary high-level expression from an individual member. A continuing puzzle, therefore, is the biological roles of the different members of these OMP multigene families. Complete genome sequences are now available for two geographically divergent strains of E. ruminantium (Caribbean and South Africa strains). Comparison of these sequences revealed amino acid sequence diversity in MAP1 (89% identity), which is known to confer protection in a mouse model and to be the multigene family member primarily expressed in mammalian cells. Surprisingly, however, the greatest sequence diversity (79% identity) was in the less-studied map1-2 gene. We investigated here whether this map1-2 diversity was a general feature of E. ruminantium in different cultured African strains and in organisms from infected sheep. Comparison of MAP1-2s revealed amino acid identities of 75 to 100% (mean of 86%), compared to 84 to 100% (mean of 89%) for MAP1s. Interestingly, MAP1-2s varied independently of MAP1s such that E. ruminantium strains with similar MAP1s had diverse MAP1-2s and vice versa. Different MAP1-2s were found in individual infected sheep. Different regions of a protein may be subjected to different evolutionary forces because of recombination and/or selection, which results in those regions not agreeing with a phylogeny deduced from the whole molecule. This appears to be true for both MAP1 and MAP1-2, where statistical likelihood methods detect heterogeneous evolutionary rates for segments of both molecules. Sera from infected cattle recognized a MAP1-2 variable-region peptide in enzyme-linked immunosorbent assay, but less strongly and consistently than a MAP1 peptide (MAP1B). Heterologous protective immunity may depend on recognition of a complex set of varying OMP epitopes.

show abstract

“…Brain biopsies were done on sheep that exhibited febrile reactions on day 3 of the reaction as described previously (39). In addition, plasma was prepared from the blood of all clinically reacting sheep on days 2 and 3 of the febrile reaction for isolation of C. ruminantium in bovine endothelial-cell culture to demonstrate transmission of heartwater by feeding ticks (3,4). A postmortem examination was done on sheep that died to confirm that death was due to heartwater, and brain crush smears were prepared and examined for C. ruminantium colonies in endothelial cells of brain capillaries (32).…”

Section: Methodsmentioning

confidence: 99%

Antibody Responses to MAP 1B and Other Cowdria ruminantium Antigens Are Down Regulated in Cattle Challenged with Tick-Transmitted Heartwater

Semu¹,

Peter²,

Mukwedeya

et al. 2001

Clin Diagn Lab Immunol

View full text Add to dashboard Cite

Serological diagnosis of heartwater orThe rickettsia Cowdria ruminantium is the causative agent of heartwater, an acute, fatal infectious disease of domestic and wild ruminants (5, 40) which is transmitted by ticks of the genus Amblyomma (45). Efforts to study the epidemiology of heartwater and to implement disease control have been hampered by the lack of reliable serodiagnostic tests. Available tests are based on cultured organisms or antigen extracts (7,9,13,15,23,26,34,36) and on the major antigenic protein of C. ruminantium, MAP 1 (2), which has been targeted as a diagnostic antigen because of its immunodominance (2,11,14). These tests are, however, limited by the extensive antigenic similarity between C. ruminantium and closely related agents of the genus Ehrlichia (1,8,12,14,17,35,41), some of which also infect ruminants. Recently, a partial fragment of MAP 1, MAP 1B, which spans amino acids 47 to 92 of the mature protein (42, 43), has been shown to have high specificity for C. ruminantium in an indirect enzyme-linked immunosorbent assay (ELISA) (24,25,43). The assay does not detect antibodies to ehrlichial agents infecting domestic ruminants, such as E. bovis, E. ovina, and E. phagocytophila. It does detect antibodies to E. canis (which infects dogs) and antibodies to E. chaffeensis (a pathogen of humans). In the United States, E. chaffeensis infects white-tailed deer (6, 16), a species that is highly susceptible to heartwater. In areas of Zimbabwe (22,43) and the Caribbean (24, 25) that are designated heartwater free by the absence of Amblyomma ticks and clinical heartwater, the MAP 1B indirect ELISA demonstrated a high specificity with cattle, sheep and goat sera. This assay is also reliable for the detection of experimental infections in small ruminants, and it detects antibodies to geographically diverse C. ruminantium isolates from different countries (24, 43). Hence, its use has been proposed for diagnosis and surveillance of heartwater.In a preliminary serological survey of heartwater in Zimbabwe using the MAP 1B indirect ELISA, only 33% of cattle sera from areas with endemic heartwater infection tested positive (22). The low seroprevalence was unexpected, given the high infection pressure in these regions and the consequent likely high prevalence of infection (27). Epidemiological studies conducted on some of these farms over several years demonstrated a tick infection rate of 10% and a vector attachment rate of between one and four ticks every 2 days (31). At this tick attack rate, it was estimated that cattle were exposed to

show abstract

In vitro isolation of Cowdria ruminantium from plasma of infected ruminants

Cited by 29 publications

References 4 publications

Detection of the Agent of Heartwater, Cowdria ruminantium , in Amblyomma Ticks by PCR: Validation and Application of the Assay to Field Ticks

Detection of the Agent of Heartwater, Cowdria ruminantium , in Amblyomma Ticks by PCR: Validation and Application of the Assay to Field Ticks

Diversity of Ehrlichia ruminantium Major Antigenic Protein 1-2 in Field Isolates and Infected Sheep

Antibody Responses to MAP 1B and Other Cowdria ruminantium Antigens Are Down Regulated in Cattle Challenged with Tick-Transmitted Heartwater

Contact Info

Product

Resources

About