Tick-borne rickettsial diseases (TBRD) are commonly encountered in medical and veterinary clinical settings. The control of these diseases is difficult, requiring disruption of a complex transmission chain involving a vertebrate host and ticks. The geographical distribution of the diseases is related to distribution of the vector, which is an indicator of risk for the population. A total of 1,107 ticks were collected by tick dragging from forests, ecotourism parks and hosts at 101 sites in 22 of the 32 states of Mexico. Collected ticks were placed in 1.5 mL cryovials containing 70% ethanol and were identified to species. Ticks were pooled according to location/host of collection, date of collection, sex, and stage of development. A total of 51 ticks were assayed by polymerase chain reaction (PCR) to confirm species identification using morphological methods. A total of 477 pools of ticks were assayed using PCR techniques for selected tick-borne pathogens. Anaplasma phagocytophilum was the most commonly detected pathogen (45 pools), followed by, Ehrlichia (E.) canis (42), Rickettsia (R.) rickettsii (11), E. chaffeensis (8), and R. amblyommii (1). Rhipicephalus sanguineus was the tick most frequently positive for selected pathogens. Overall, our results indicate that potential tick vectors positive for rickettsial pathogens are distributed throughout the area surveyed in Mexico.
Anaplasma phagocytophilum ; Animals* ; Ehrlichia ; Ehrlichia canis ; Ehrlichia chaffeensis ; Ethanol ; Forests ; Humans* ; Mexico* ; Polymerase Chain Reaction ; Rhipicephalus sanguineus ; Rickettsia ; Ticks* ; Vertebrates

The study of canine vector-borne diseases in the Philippines started in the 1970s but only gained interest in the past decade. Characterization of such diseases in the Philippines remains incomplete, thus, it is necessary to obtain additional information on the prevalence and diversity of canine tick-borne diseases in the country. In this study, blood samples were obtained at two veterinary clinics in Metro Manila, Philippines from 114 dogs suspected of having canine tick-borne pathogens. Polymerase chain reaction (PCR) was performed on whole blood DNA extracts followed by sequencing, and the following pathogens were detected: Hepatozoon (H.) canis (5.26%), Babesia (B.) vogeli (5.26%), Ehrlichia (E.) canis (4.39%), and Anaplasma platys (3.51%). Additionally, a set of multiplex PCR primers were developed to detect H. canis, Babesia spp. (B. canis and B. vogeli), and E. canis in canine blood. Multiplex and conventional single-reaction PCR results for the 114 dog blood samples were similar, except for one H. canis sample. Multiplex PCR is, therefore, a useful tool in screening infected dogs in veterinary clinics. This study's results, together with those of previous studies in the country, show that canine vector-borne pathogens are an emerging veterinary concern in the Philippines.
Anaplasma* ; Animals ; Babesia* ; DNA ; Dogs* ; Ehrlichia canis* ; Ehrlichia* ; Hospitals, Animal ; Mass Screening ; Multiplex Polymerase Chain Reaction ; Philippines* ; Polymerase Chain Reaction ; Prevalence ; Tick-Borne Diseases

A total of 1,395 Haemaphysalis longicornis ticks collected from Jeju Island of Korea were examined by 16S rRNA gene-based nested PCR for the presence of infection with Anaplasma and Ehrlichia species. Template DNAs to detect the tick-borne pathogens were prepared from a total 506 tick pools. Eight genera of Anaplasma and six Ehrlichia by 16S rRNA gene PCR and sequencing analysis were identified. A. phagocytophilum was the most prevalent (27 [1.9%]) by nested PCR, followed by A. bovis (5 [0.4%]), E. chaffeensis (4 [0.2%]), and A. centrale (1 [0.1%]). In the phylogenetic analysis based on 16S rRNA sequences, eight genera of Anaplasma group (> 99.4% homology) and six Ehrlichia group (> 99.5% homology) were close to deposited A. marginale strains (AF309867, AF414874, and FJ226454) and Ehrlichia sp. (DQ324547), respectively. Three Anaplasma species groups A. phagocytophilum (group A), A. bovis (group B), and A. centrale (group C) and one Ehrlichia species E. chaffeensis (group D) were determined by comparing with Anaplasma and Ehrlichia related sequences. First, twenty-eight A. phagocytophilum clones belonging to group A were divided into 7 genotypes. The sequence similarity among genotypes A1 to A4 was very high (> 99.6%). Genotype B2 was close to A. bovis from Korea (99.7%). Genotype D1 was close to known E. chaffeensis strains (M73222, AF147752, and AY350424) and their similarity value was 99.7%. In conclusion, the genera of Anaplasma/Ehrlichia, A. phagocytophilum, and E. chaffeensis identified in predominant H. longicornis ticks were ubiquitous throughout the Jeju Island. The various native groups have been found through sequence identities and phylogenetic analysis.
Anaplasma ; Anaplasma phagocytophilum ; Clone Cells ; DNA ; Ehrlichia ; Ehrlichia chaffeensis ; Genes, rRNA ; Genotype ; Korea ; Polymerase Chain Reaction ; Ticks

Ehrlichia (E.) canis is a Gram-negative obligate intracellular bacterium responsible for canine monocytic ehrlichiosis. Currently, the genetic diversity of E. canis strains worldwide is poorly defined. In the present study, sequence analysis of the nearly full-length 16S rDNA (1,620 bp) and the complete coding region (4,269 bp) of the gp200 gene, which encodes the largest major immunoreactive protein in E. canis, from 17 Taiwanese samples was conducted. The resultant 16S rDNA sequences were found to be identical to each other and have very high homology (99.4~100%) with previously reported E. canis sequences. Additionally, phylogenetic analysis of gp200 demonstrated that the E. canis Taiwanese genotype was genetically distinct from other reported isolates obtained from the United States, Brazil, and Israel, and that it formed a separate clade. Remarkable variations unique to the Taiwanese genotype were found throughout the deduced amino acid sequence of gp200, including 15 substitutions occurring in two of five known species-specific epitopes. The gp200 amino acid sequences of the Taiwanese genotype bore 94.4~94.6 identities with those of the isolates from the United States and Brazil, and 93.7% homology with that of the Israeli isolate. Taken together, these results suggest that the Taiwanese genotype represents a novel strain of E. canis that has not yet been characterized.
Amino Acid Sequence ; Animals ; Bacterial Proteins/chemistry/*genetics ; Dogs ; Ehrlichia canis/*classification/*genetics ; Genotype ; Molecular Sequence Data ; *Phylogeny ; RNA, Ribosomal, 16S/genetics ; Sequence Alignment ; Sequence Analysis, Protein ; Taiwan

Ehrlichia ; Ehrlichiosis ; microbiology

Amplification of the 16S rRNA gene from a blood sample obtained from a dog in southeastern Brazil was used to confirm a naturally acquired Ehrlichia (E.) canis infection. Following isolation and culturing of the new bacterial strain called Uberlandia, partial sequences of the dsb and p28 genes were obtained. The dsb partial sequence of the novel strain was 100% similar to dsb gene sequences of E. canis obtained from different geographic areas around the world. Conversely, the p28 partial sequence for the E. canis Uberlandia strain differed at several nucleotides from other sequences available in GenBank. To confirm the antigenic profile of the Uberlandia strain, an indirect immunofluorescence assay against E. canis antigens was performed using dog sera collected from two different areas in Brazil (Uberlandia and Sao Paulo). The results suggest that both antigens were able to identify animals seropositive for E. canis in Brazil since these Brazilian strains appear to be highly conserved.
Animals ; Antigens, Bacterial/blood/*diagnostic use ; Bacterial Outer Membrane Proteins/genetics/metabolism ; Bacterial Proteins/*genetics/metabolism ; Base Sequence ; Brazil ; Dog Diseases/diagnosis/*microbiology ; Dogs ; Ehrlichia canis/*genetics/*immunology/isolation & purification ; Ehrlichiosis/diagnosis/microbiology/*veterinary ; Fluorescent Antibody Technique, Indirect/veterinary ; Male ; Molecular Sequence Data ; Polymerase Chain Reaction/veterinary ; RNA, Ribosomal, 16S/genetics/metabolism ; Sequence Alignment/veterinary

OBJECTIVETo evaluate the Anaplasma phagocytophilum (A. phagocytophilum), Ehrlichia canis (E. canis), Dirofilaria immitis (D. immitis) (canine heartworm), Borrelia burgdorferi (B. burgdorferi) infections in countryside dogs from Yunnan, Hainan and Anhui provinces.

METHODSSerum samples were collected from 26 dogs in Yunnan, Hainan and Anhui provinces. The samples were tested using a commercial ELISA rapid diagnostic assay kit (SNAP(®) 4Dx(®); IDEXX Laboratories, Inc. U.S.A.). Meanwhile, indirect immunofluorescence assay (IFA) recommended by WHO was conducted to detect IgG to A. phagocytophilum. Two methods were analyzed and compared.

RESULTSThe number of serologically positive dogs for IgG to A. phagocytophilum was only 2 which was from Hainan province and none of the 26 dogs responded positive for E. canis, D. immitis (canine heartworm), and B. burgdorferi by ELISA rapid diagnostic method. The number of serologically positive dogs for IgG to A. phagocytophilum was 13 (50%) by IFA method. Data of the two methods were analyzed by statistical software and the difference was statistically significant (P=0.002).

CONCLUSIONSIt can be concluded that IFA method was more sensitive than ELISA rapid diagnostic method. However, we need conduct further and intensive epidemiology survey on tick-born diseases pathogens including A. phagocytophilum, E. canis, D. immitis (canine heartworm), and B. burgdorferi which have public health significance.

Anaplasma phagocytophilum ; immunology ; Animals ; Borrelia burgdorferi ; immunology ; China ; epidemiology ; Dirofilaria immitis ; immunology ; Dirofilariasis ; blood ; epidemiology ; immunology ; Disease Vectors ; Dog Diseases ; epidemiology ; Dogs ; Ehrlichia canis ; immunology ; Ehrlichiosis ; blood ; epidemiology ; immunology ; Enzyme-Linked Immunosorbent Assay ; methods ; Fluorescent Antibody Technique, Indirect ; methods ; Immunoglobulin G ; blood ; Lyme Disease ; blood ; epidemiology ; immunology ; Tick-Borne Diseases ; epidemiology

Animals ; Ehrlichia/isolation & purification* ; Ehrlichiosis/veterinary* ; Prevalence ; Rodent Diseases/microbiology* ; Rodentia ; Species Specificity

Genomic DNAs extracted from 1,288 Haemaphysalis longicornis ticks collected from grass vegetation and various animals from nine provinces of Korea were subjected to screening by genus-specific (Ehrlichia spp. or Anaplasma spp.) real-time TaqMan PCR and speciesspecific (E. chaffeensis) nested-PCR based on amplification of 16S rRNA gene fragments. In all, 611 (47.4%) ticks tested positive for genus-specific amplification of 116 bp fragment of 16S rRNA of Ehrlichia spp. or Anaplasma spp. Subsequently, 396 bp E. chaffeensis-specific fragment of 16S rRNA was amplified from 4.2% (26/611) tick samples. The comparison of the nucleotide sequence of 16S rRNA gene from one tick (EC-PGHL, GeneBank accession number AY35042) with the sequences of 20 E. chaffeensis strains available in the database showed that EC-PGHL was 100% identical or similar to the Arkansas (AF416764), the Sapulpa (U60476) and the 91HE17 (U23503) strains. The phylogenetic analysis also revealed that the E. chaffeensis EC-PGHL formed a single cluster with the above strains. This is the first study to report molecular detection and phylogenetic analysis of E. chaffeensis from H. longicornis ticks in Korea. The implicit significance of E. chaffeensis infection in H. longicornis ticks in Korea is discussed.
Anaplasma/growth&development ; Animals ; Base Sequence ; Cloning, Molecular ; DNA, Bacterial/chemistry/genetics ; Ehrlichia chaffeensis/*genetics/growth&development ; Ehrlichiosis/*epidemiology/microbiology ; Korea/epidemiology ; Molecular Sequence Data ; Phylogeny ; Polymerase Chain Reaction ; Prevalence ; RNA, Ribosomal, 16S/chemistry/genetics ; Sequence Alignment ; Sequence Analysis, DNA ; Ticks/*microbiology

OBJECTIVETo provide further pathogenic evidence of Granulocytic ehrlichia infection in China.

METHODSSpecific primers derived from 444-Epank gene were used to amplify Granulocytic ehrlichia DNA from specimens of ticks, animals and human blood. PCR products of ticks were cloned and sequenced.

RESULTS444 bp specific DNA fragments were amplified from 2 of 62 pools of Ixodes persulcatus collected from Heilongjiang province and 1 of 129 blood specimens from forest workers in Inner Mongolia. Eight animal specimens were negative. PCR products from ticks were then cloned and sequenced. It differed at 23 positions in comparison to American strain (AF047897) with 94.9% homology. The homology of deduced ammonia was 88.44%.

CONCLUSIONOur findings further confirmed that Granulocytic ehrlichia infection did exist in China.

DNA, Bacterial ; analysis ; Ehrlichia ; classification ; genetics ; isolation & purification ; Ehrlichiosis ; microbiology ; Genes, Bacterial ; Humans ; Phylogeny ; Polymerase Chain Reaction ; Sequence Analysis, DNA