Molecular occurrence and genetic diversity of Ehrlichia canis in naturally infected dogs from Thailand.

Canine monocytic ehrlichiosis is cause by Ehrlichia canis resulting in hematologic disorders and severe clinical signs. The aim of this study was to scrutinize the molecular detection and genetic diversity of E. canis based on the trp36 gene in dogs from Thailand's northern and central regions. A total of 120 dogs blood samples were amplified for trp36 gene of E. canis using the polymerase chain reaction (PCR). Forty-seven out of 120 dog blood samples (39.16%, 47/120) were positive for E. canis the trp36 DNA with 790 bp of PCR amplicon size. The factor significantly associated with E. canis infection is animal housing status (p < 0.05). Sequence and phylogenetic analysis showed that E. canis trp36 gene of Thailand isolates was clustered into 1st clade with similarity ranging from 95.65 to 100% together with the US genogroup. The 14 haplotypes of the trp36 gene shown in TCS network exhibited that haplotype #1-4 was found in Thailand. The entropy analysis of the trp36 gene illustrated 751 polymorphic sites and 271 entropy peaks of nucleic and amino acid sequences, respectively. Hence, these findings are crucial for better understanding the epidemiology of Ehrlichia infection and could be helpful for implementing control measures in Thailand.

Ehrlichia canis: Molecular characterization and genetic diversity based on the p28 and trp36 genes.

Ehrlichia canis is a common tick-borne intracellular pathogen causing canine monocytic ehrlichiosis (CME) in dogs worldwide. The aims of this study were to investigate the genetic diversity and antigenicity of E. canis based on the p28 and trp36 genes in dogs in Thailand. The E. canis p28 and trp36 genes were amplified by the polymerase chain reaction (PCR) and cloned for sequencing and bioinformatic analyses. 36% (44/120) of dog blood samples were positive for E. canis DNA consisting of p28 (31%, 14/44) and trp36 (69%, 30/44) genes with 792 and 882 bp of PCR products size, respectively. The E. canis TRP36 from all Thailand sequences exhibited encoded nine amino acids (TEDSVSAPA) with 11 copies of tandem repeats along the sequences. The phylogenetic trees of E. canis, using the p28 and trp36 genes, exhibited that the Thailand isolates fell into two clades and one clade with similarity ranging from 55.95 to 100% and 100%, respectively. The results of diversity analysis revealed 10 and 20 haplotypes of the p28 and trp 36 genes, respectively. The entropy analysis of the p28 and trp36 nucleic acid sequences showed 442 and 1321 high entropy peaks respectively, whereas those of the P28 and TRP36 amino acid sequences showed 477 and 388 high entropy peaks, respectively. For B-cell epitopes analysis, the conserved amino acid of P28 and TRP36 sequences has been also demonstrated. Therefore, the results could be utilized to improve the understanding of phylogenetic relationship, genetic diversity and antigenicity of E. canis Thailand isolates.

First study on molecular detection of three major canine tick-borne pathogens in subclinically infected dogs in Chiang Mai, Thailand.

Background and Aim: Canine tick-borne pathogens (CTBPs) are an important cause of morbidity in dogs in Thailand. This study aimed to evaluate the occurrence of three CTBPs in clinically normal, owned dogs to understand the risk for the general canine population. We also examined sex, age, tick infestation, and packed cell volume (PCV) of the animals in association with active infection of the CTBPs. Materials and Methods: A total of 139 dogs were included in the study. Blood samples were collected for thin blood smear, PCV and nested polymerase chain reaction (PCR) assay. Statistical analyses were performed to examine the association between individual factors and CTBP infection status determined by PCR. In addition, sensitivity, specificity, and Cohen's kappa were calculated to assess the utility of routine blood smear. Results: The PCR results showed that 31 dogs (22.3%) were infected with at least one of the three pathogens. The occurrence rate for Ehrlichia canis, Anaplasma platys, and Hepatozoon canis was 2.2% (3/139), 18.7% (24/139), and 2.8% (4/139), respectively. There were two cases of coinfection with A. platys and E. canis. The univariate analyses did not yield any associations between recorded variables and the active infection. Microscopic examination showed good sensitivity and agreement only for H. canis (Sn: 75%, 95% confidence interval: 24.9-98.7, k=0.85). Conclusion: Our findings confirmed the endemicity of the CTBPs in owned canine population in the study site. In-depth epidemiological investigation would be warranted to elucidate environmental risk factors for CTBP infection.

Molecular discrimination and genetic diversity of three common tick-borne pathogens in dogs in Thailand.

There was little information regarding the occurrence of canine vector-borne disease (CVBDs) in shelter dogs in Thailand. This work is the first report regarding a molecular method used to determine the occurrence and genetic diversity of three canine tick-borne pathogens (TBPs) (Hepatozoon canis, Anaplasma platys and Ehrlichia canis) in blood samples from 275 shelter dogs in the north and central areas of Thailand. The PCR results based on the 18S rRNA and 16S rRNA genes showed that 71 (25.82%) dogs were positive for at least a TBP. The overall occurrence rates of H. canis, A. platys and E. canis infections were 1.81, 16.36 and 7.64%, respectively. For the phylogenetic analysis, A. platys 16S rRNA gene was genetically diverse, while H. canis 18S rRNA and E. canis 16S rRNA genes were conserved. The haplotype diversity exhibited 12 and 2 haplotypes as well as 78 and 178 polymorphic sites of A. platys and E. canis 16S rRNA genes, respectively. Our findings could be used to improve the understanding of phylogeny and genetic diversity of TBP rRNA genes and used to ameliorate the diagnosis and control programmes for the diseases in Thailand.