Serological evidence of Ehrlichia minasensis infection in Brazilian dogs.

Ehrlichia spp. are important tick-borne pathogens of animals in Brazil, and Ehrlichia canis is the most prevalent species infecting dogs. Moreover, Ehrlichia minasensis has also recently been identified as a novel ehrlichial agent that infects cattle in Brazil. The objective of this study was to determine whether dogs could be infected by E. minasensis. To investigate this possibility, sera (n = 429) collected from dogs in the Pantanal region were retrospectively analyzed for the presence of antibodies against E. canis and E. minasensis. Canine sera were screened by two isolates of E. canis in indirect immunofluorescence assay (IFA) and the majority (n = 298; 69.4%) had antibodies with endpoint titers ranging from 80 to 327,680. In order to further confirm E. canis-specific antibodies, IFA positive sera were analyzed by ELISA using E. canis-specific peptides (i.e. TRP19 and TRP36 US/BR/CR), which detected E. canis antibodies in 80.2% (239/298) of the dog sera. Fifty-nine (13.7%) samples had detectable antibodies to E. canis by IFA but were negative by E. canis peptide ELISA. These sera were then tested by E. minasensis IFA (Cuiaba strain) as antigen and 67.8% (40/59) were positive (titers ranging from 80 to 20,480). Eleven sera had antibody titers against E. minasensis at least two-fold higher than observed for E. canis and suggests that these dogs were previously infected with E. minasensis. The results of the present study suggest that multiple ehrlichial agents infect dogs in Brazil, which highlights the need to consider different Ehrlichia spp. in Brazilian dogs, particularly in areas where dogs are frequently exposed to multiple tick species. This investigation is the first to provide serologic evidence of E. minasensis infection in dogs from Brazil.

Genetic and antigenic diversities of major immunoreactive proteins in globally distributed Ehrlichia canis strains.

The extent of knowledge regarding the diversity of globally distributed Ehrlichia canis strains has been limited to information gained from a few evolutionarily conserved genes. In this study, E. canis strains from the United States (strain Jake [US]), Brazil (strain São Paulo [BR]), and Israel (strain 611 [IS] and Ranana [IS-R]) were used to examine the antigenic and genetic diversities of four well-characterized major immunoreactive protein genes/proteins. gp36 and gp200 were the most divergent genes, and nucleotide substitutions in the gp36 tandem repeat region of the IS strain, but not the IS-R strain, resulted in two amino acid differences (S-->P and P-->T) in each nine-amino-acid repeat (epitope-containing region). DNA sequences of gp19 and gp140 were completely conserved in the US and BR strains, but differences were found in the Israeli strains, including two fewer tandem repeats in gp140 and a single amino acid substitution in gp19 from the IS strain. E. canis whole-cell lysates from each isolate were examined by Western immunoblotting using sera from naturally infected dogs from each country, and four major immunoreactive proteins (gp19, gp36, gp140, and gp200) were identified in each strain using protein-specific antisera. The US and BR strains exhibited highly conserved immunoreactive protein profiles, while some differences were identified in the IS strain. Sera from naturally infected Israeli dogs confirmed gene sequencing information, which demonstrated two distinct E. canis strains, defined by the gp36 gene. Conversely, gp19 was strongly reactive and present in all E. canis isolates. gp140 and gp200 were also present in all strains, although gp140 in the IS strain had two fewer tandem repeats and exhibited a smaller mass.