Facultad de medicina veterinaria, Grupo de Investigación en Ciencias Veterinarias, Centauro, Universidad de Antioquia, Calle 70 No. 52-21, Medellín, Colombia; Department of Pathology, Center for Biodefense and Emerging Infectious Diseases, Sealy Institute for Vaccine Development, University of Texas Medical Branch, 301 University Boulevard, Galveston, TX, 77555-0609, USA.
Facultad de medicina veterinaria, Grupo de Investigación en Ciencias Veterinarias, Centauro, Universidad de Antioquia, Calle 70 No. 52-21, Medellín, Colombia.
Ticks Tick Borne Dis. 2020 May;11(3):101367. doi: 10.1016/j.ttbdis.2019.101367. Epub 2020 Jan 7.
Ehrlichia canis is the etiologic agent of a highly prevalent tick-borne disease, canine monocytic ehrlichiosis (CME). Four defined E. canis genotypes based on the trp36 gene sequences have been reported, three of them identified in North or South America. The diversity of E. canis has been investigated using genetic and serologic approaches based on distinct 36 kDa tandem repeat protein (trp36) gene sequences that have been reported. The main objectives of this study were to determine the prevalence of E. canis infection in dogs from Medellín, Colombia by PCR and determine the E. canis diversity using molecular and serologic approaches. Blood was collected from dogs (n = 300) with clinical signs of CME for PCR detection of E. canis 16S rRNA, dsb and trp36 DNA. Phylogenetic analysis of trp36 gene sequences was performed using MEGA. A serological evaluation was performed using immunofluorescence microscopy and ELISA with species-specific peptides from E. canis TRP19 and TRP36 (3 genotypes) and E. chaffeensis (TRP32). E. canis DNA (16S rRNA and/or dsb) was detected in 18 % (53/300) of dogs by PCR amplification. The trp36 gene was amplified and sequenced from 35/53 16S rRNA/dsb PCR positive samples revealing three genotypes: United States (US; n = 21), Costa Rica (CR; n = 11), and Brazil (BR; n = 3). Most dogs (33/35) with detectable trp36 DNA had anti-E. canis TRP19 and TRP36 peptide antibodies that corresponded to the genotype detected by PCR. Dogs that had antibodies to the TRP19 peptide (82/300; 38 %), also had antibodies to one or more genotype-specific TRP36 peptides. Based on TRP36 serology, the dogs exhibited highest frequency of infection with the US genogroup (US = 26), followed by the CR genogroup (CR = 19) and the BR genogroup (BR = 11). Notably, 26/53 trp36 PCR positive dogs had detectable antibodies to multiple E. canis genotypes (US/BR/CR = 8, BR/CR = 7, US/CR = 6 and US/BR = 5) suggesting coinfection or multiple sequential infections with different genotypes. Colombian dogs did not have antibodies to E. chaffeensis as determined by a TRP32 species-specific ELISA. Our results demonstrate the presence of three previously defined genotypes in North and South America in Colombian dogs (US, BR, CR). These results also demonstrate that TRP19 and TRP36 serology can provide valuable information regarding E. canis exposure and the potential genotype(s) involved in infection.
犬埃立克体是一种高度流行的蜱传疾病——犬单核细胞埃立克体病（CME）的病原体。根据 trp36 基因序列，已经报道了四种确定的犬埃立克体基因型，其中三种在北美或南美被发现。已经使用基于不同 36 kDa 串联重复蛋白（trp36）基因序列的遗传和血清学方法研究了犬埃立克体的多样性，这些序列已被报道。本研究的主要目的是通过 PCR 检测来自哥伦比亚麦德林的犬感染犬埃立克体的流行情况，并通过分子和血清学方法确定犬埃立克体的多样性。从有 CME 临床症状的犬（n=300）采集血液，进行 PCR 检测犬埃立克体 16S rRNA、dsb 和 trp36 DNA。使用 MEGA 进行 trp36 基因序列的系统发育分析。使用针对犬埃立克体 TRP19 和 TRP36（3 种基因型）和犬埃立克体 chaffeensis（TRP32）的种特异性肽的免疫荧光显微镜和 ELISA 进行血清学评估。通过 PCR 扩增在 18%（53/300）的犬中检测到犬埃立克体 DNA（16S rRNA 和/或 dsb）。从 53 个 16S rRNA/dsb PCR 阳性样本中扩增和测序 trp36 基因，揭示了三种基因型：美国（US；n=21）、哥斯达黎加（CR；n=11）和巴西（BR；n=3）。大多数（33/35）可检测到 trp36 DNA 的犬具有抗犬埃立克体 TRP19 和 TRP36 肽抗体，与 PCR 检测到的基因型相对应。具有 TRP19 肽抗体的犬（82/300；38%）也具有一种或多种基因型特异性 TRP36 肽抗体。基于 TRP36 血清学，狗表现出最高的美国基因型（US=26）感染频率，其次是哥斯达黎加基因型（CR=19）和巴西基因型（BR=11）。值得注意的是，26/53 trp36 PCR 阳性犬具有可检测到的针对多种犬埃立克体基因型（US/BR/CR=8、BR/CR=7、US/CR=6 和 US/BR=5）的抗体，表明存在合并感染或多种基因型的连续感染。通过 TRP32 种特异性 ELISA 确定哥伦比亚犬没有针对埃立克体 chaffeensis 的抗体。我们的结果表明，在哥伦比亚犬中存在三种以前在北美和南美定义的基因型（美国、巴西、哥斯达黎加）。这些结果还表明，TRP19 和 TRP36 血清学可以提供有关犬埃立克体暴露和潜在基因型（s）参与感染的有价值信息。
