Prevalence of Anaplasma phagocytophilum in Ixodes ricinus ticks determined by polymerase chain reaction with two pairs of primers detecting 16S rRNA and ankA genes.

A total of 684 Ixodes ricinus ticks (321 nymphs, 184 males, and 179 females) were collected by flagging lower vegetation in 6 forest districts located on the territory of Lublin province (eastern Poland). Ticks were examined by polymerase chain reaction (PCR) method for the presence of Anaplasma phagocytophilum DNA with two pairs of primers: EHR521/EHR747 for detecting 16S rRNA gene, and LA6/LA1 for detecting ankA gene. To study the relationship between infection in ticks and people occupationally exposed to tick bite, blood serum samples of 261 forestry workers employed in the same forest districts were examined by immunofluorescence method for the presence of specific antibodies against A. phagocytophilum. A total of 70 ticks out of 684 examined (10.2%) showed the presence of A. phagocytophilum 16S rRNA gene. The prevalence of infection was significantly dependent on tick's stage (chi-square=49.2, p<0.00001) and geographical locality (chi-square=34.4, p<0.00001). The percentage of I. ricinus females infected with A. phagocytophilum (24.6%) was significantly greater compared to males (6.5%) and nymphs (4.4%) (p<0.00001). Only 19 ticks out of 684 examined (2.8%) showed the presence of A. phagocytophilum ankA gene, significantly less compared to 16S rRNA gene (p<0.00001). The prevalence of infection demonstrated by the presence of ankA gene was also significantly dependent on tick's stage (chi-square=23.6, p<0.00001) but not on locality (chi-square=9.8, p=0.082). A significant correlation was found between the presence of A. phagocytophilum 16S rRNA gene in I. ricinus female ticks from the particular forest districts and the serologic response to A. phagocytophilum of forestry workers employed in the same districts (p<0.05). No significant correlation was found between the presence of A. phagocytophilum ankA gene in I. ricinus ticks and serologic response of exposed workers. In conclusion, detection of A. phagocytophilum infection in ticks by PCR with the use of EHR521/EHR747 primers detecting 16S rRNA gene is significantly more sensitive compared to LA6/LA1 primers detecting ankA gene.