Detection and quantification of Anaplasma phagocytophilum and Babesia spp. in Ixodes ricinus ticks from urban and rural environment, northern Poland, by real-time polymerase chain reaction

Abstract

Anaplasma phagocytophilum and Babesia spp. are emerging tick-borne pathogens which can threaten human health. A duplex real-time PCR and qPCRs with primers and probes targeting 97 and 116 bp fragments of 16S rRNA and 18S rRNA genes, respectively, were used for qualitative and quantitative detection of both pathogens in Ixodes ricinus ticks. Altogether 1875 ticks (1084 adults and 791 nymphs) were collected from rural and urban habitats in northern Poland. Of them, at least 0.9 % were found to be infected with A. phagocytophilum while 2.5 % with Babesia spp. A comparison of the infection rates by the tick stage, the type of area, the collection site, habitats of different tick density and by the month of collection was done. The prevalence of pathogens was significantly lower in nymphs than in adult ticks (p = 0.02) and in rural areas than in urban areas (p = 0.007). Four different 16S rRNA gene variants of A. phagocytophilum were determine, however none of them showed 100 % identity with compared sequences isolated from human patients. The dominant Babesia species was B. venatorum. Results of qPCRs with circular and linearized forms of plasmids used as the standards showed significant difference in the pathogen loads (p = 0.001). The copy numbers of A. phagocytophilum and Babesia spp. estimated from the linear plasmids were 28.7 and 5.1 times lower, respectively, when compared with their circular forms, and were accepted as more reliable. The average number of copies of 16S rRNA gene of A. phagocytophilum in the positive I. ricinus samples were 3.39 × 105 ± 6.09 × 105. The mean copy number of 18S rRNA gene of Babesia spp. was ~2.55 × 105 ± 1.04 × 106. We confirmed the presence of A. phagocytophilum and Babesia spp. in I. ricinus in both rural and urban environments. The determined low infection rates suggests, however, that the risk for local population and tourists to acquire infection is also low. Moreover, we confirmed recent findings that serious overestimation by circular plasmid DNA makes it less suitable as a standard and that the linear standards should be recommended for qPCR.

Babesia spp. in ticks and wildlife in different habitat types of Slovakia

“…Importantly, we report here for the first time B. canis infection of I. ricinus from Slovakia. Thus, we confirm the previous conjecture of the infection of I. ricinus with B. canis as noted in a study from Poland [ 67 , 68 ]. However, our finding does not allow us to draw conclusions on the competence of I. ricinus as a vector of this parasite.…”

Detection of tick-borne pathogens in questing Ixodes ricinus in the French Pyrenees and first identification of Rickettsia monacensis in France

“…In our study, Babesia spp. prevalence was 0.4% which is in accordance with similar studies carried in European countries such as Poland, Germany, Slovakia and Switzerland where infection rates were between 0.4 and 4.5% [ 16 , 44 ]. In fact, I. ricinus carrier rates of Babesia spp.…”

Comparison of tick-borne pathogen prevalence in Ixodes ricinus ticks collected in urban areas of Europe

“…The analysis showed a greater risk of A. phagocytophilum infections in Eastern Europe than in Western Europe. Similar infection rates in I. ricinus ticks were reported in studies from the Czech Republic (0.8-7.2%) 17,51 , Hungary (8.8%) 11 , Poland (1.7-14.0%) 1,43,44 , Slovakia (2.9-7.2%) 29,46,57 and the Ukraine (5.2%) 58 . Significantly lower infection rates were observed in Western Europe, e.g., Switzerland (1.4-1.5%) 18,27 and Germany (1.8-4.4%) 19,[23][24][25]35,50 .…”