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Ehrlichia ruminantium major antigenic protein gene (map1) variants are not geographically constrained and show no evidence of having evolved under positive selection pressure.

ABSTRACT: In a search for tools to distinguish antigenic variants of Ehrlichia ruminantium, we sequenced the major antigenic protein genes (map1 genes) of 21 different isolates and found that the sequence polymorphisms were too great to permit the design of probes which could be used as markers for immunogenicity. Phylogenetic comparison of the 21 deduced MAP1 sequences plus another 9 sequences which had been previously published did not reveal any geographic clustering among the isolates. Maximum likelihood analysis of codon and amino acid changes over the phylogeny provided no statistical evidence that the gene is under positive selection pressure, suggesting that it may not be important for the evasion of host immune responses.

SUBMITTER: Allsopp MT

PROVIDER: S-EPMC88518 | biostudies-literature | 2001 Nov

REPOSITORIES: biostudies-literature

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Ehrlichia ruminantium major antigenic protein gene (map1) variants are not geographically constrained and show no evidence of having evolved under positive selection pressure.

Allsopp M T MT Dorfling C M CM Maillard J C JC Bensaid A A Haydon D T DT van Heerden H H Allsopp B A BA

Journal of clinical microbiology 20011101 11

In a search for tools to distinguish antigenic variants of Ehrlichia ruminantium, we sequenced the major antigenic protein genes (map1 genes) of 21 different isolates and found that the sequence polymorphisms were too great to permit the design of probes which could be used as markers for immunogenicity. Phylogenetic comparison of the 21 deduced MAP1 sequences plus another 9 sequences which had been previously published did not reveal any geographic clustering among the isolates. Maximum likelih ...[more]

PMID: 11682561

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Project description:The obligate intracellular pathogenic bacterium, Ehrlichia ruminantium, is the causal agent of heartwater, a fatal disease in ruminants transmitted by Amblyomma ticks. So far, three strains have been attenuated by successive passages in mammalian cells. The attenuated strains have improved capacity for growth in vitro, whereas they induced limited clinical signs in vivo and conferred strong protection against homologous challenge. However, the mechanisms of pathogenesis and attenuation remain unknown. In order to improve knowledge of E. ruminantium pathogenesis, we performed a comparative transcriptomic analysis of two distant strains of E. ruminantium, Gardel and Senegal, and their corresponding attenuated strains. Overall, our results showed an upregulation of gene expression encoding for the metabolism pathway in the attenuated strains compared to the virulent strains, which can probably be associated with higher in vitro replicative activity and a better fitness to the host cells. We also observed a significant differential expression of membrane protein-encoding genes between the virulent and attenuated strains. A major downregulation of map1-related genes was observed for the two attenuated strains, whereas upregulation of genes encoding for hypothetical membrane proteins was observed for the four strains. Moreover, CDS_05140, which encodes for a putative porin, displays the highest gene expression in both attenuated strains. For the attenuated strains, the significant downregulation of map1-related gene expression and upregulation of genes encoding other membrane proteins could be important in the implementation of efficient immune responses after vaccination with attenuated vaccines. Moreover, this study revealed an upregulation of gene expression for 8 genes encoding components of Type IV secretion system and 3 potential effectors, mainly in the virulent Gardel strain. Our transcriptomic study, supported by previous proteomic studies, provides and also confirms new information regarding the characterization of genes involved in E. ruminantium virulence and attenuation mechanisms.

Project description: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.

Project description:BACKGROUND: The rickettsial bacterium Ehrlichia ruminantium is the causative agent of heartwater in ruminants. A better understanding of the population genetics of its different strains is, however, needed for the development of novel diagnostic tools, therapeutics and prevention strategies. Specifically, the development of effective vaccination policies relies on the proper genotyping and characterisation of field isolates. Although multi-locus sequence typing (MLST) has been recently developed, only strains from geographically restricted collections have been analysed so far. The expansion of the MLST database to include global strains with different geographic origins is therefore essential. In this study, we used a panel of reference strains from geographically diverse origins and field samples of E. ruminantium detected from its vector, Amblyomma variegatum, in heartwater-endemic areas in Uganda. RESULTS: A total of 31 novel alleles (six, four, six, three, two, five, three, and two for gltA, groEL, lepA, lipA, lipB, secY, sodB, and sucA loci, respectively) and 19 novel sequence types (STs) were identified. Both neighbour-joining and minimum spanning tree analyses indicated a high degree of genetic heterogeneity among these strains. No association was observed between genotypes and geographic origins, except for four STs from West African countries. When we performed six different tests for recombination (GeneConv, Bootscan, MaxChi, Chimaera, SiScan, and 3Seq) on concatenated sequences, four possible recombination events were identified in six different STs. All the recombination breakpoints were located near gene borders, indicating the occurrence of intergenic recombination. All four STs that localized to a distinct group in clustering analysis showed evidence of identical recombination events, suggesting that recombination may play a significant role in the diversification of E. ruminantium. CONCLUSIONS: The compilation of MLST data set across the African continent will be particularly valuable for the understanding of the existing genetic diversity of field isolates in African countries. Comprehensive information on the degree of cross-protection between strains and further understanding of possible relationships between genotypes and phenotypes such as vaccine efficacy are expected to lead to the development of region-specific vaccination strategies.

Dataset Information

Ehrlichia ruminantium major antigenic protein gene (map1) variants are not geographically constrained and show no evidence of having evolved under positive selection pressure.

Publications

Ehrlichia ruminantium major antigenic protein gene (map1) variants are not geographically constrained and show no evidence of having evolved under positive selection pressure.

Similar Datasets

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