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Table 1 PCR methods, target genes, primers, expected PCR product sizes, and reaction and amplification conditions as well as references used for detection of tick-borne pathogens

From: Molecular investigation of bacterial and protozoal pathogens in ticks collected from different hosts in Turkey

Organism Molecular diagnostic method Target gene Primer PCR product size (bp) Reaction content Reaction conditions References
Hepatozoon spp. Conventional PCR 18S rDNA 5′-ATACATGAGCAAAATCTCAAC-3′
5′-CTTATTATTCCATGCTGCAG-3′
650 25 µl reaction volume (12.5 μl PCR master mix, 1 μl primers, 2 μl DNA) 95 °C 5 min, 34× (95 °C 30 s, 53 °C 30 s, 72 °C 90 s), 72 °C 5 min [11, 55]
Theileria spp.
Babesia spp.
Nested PCR 1 18S rDNA 5′-GTGAAACTGCGAATGGCTCATTAC-3′
5-AAGTGATAAGGTTCACAAAACTTCCC-3
1609–1523 25 µl reaction volume (12.5 μl PCR master mix, 1 μl primers, 1 μl DNA) 94 °C 1 min, 30× (98 °C 10 s, 55 °C 15 s, 68 °C 45 s), 68 °C 5 min [10]
Nested PCR 2 5′-GGCTCATTACAACAGTTATAGTTTATTTG-3′
5′-CGGTCCGAATAATTCACCGGAT-3′
1544–1454
Anaplasma spp. Nested PCR 1 Msp4 5′-ATGAATTACAGAGAATTGCTTGTAGG-3′
5-TTAATTGAAAGCAAATCTTGCTCCTATG-3′
849 25 µl reaction volume (12.5 μl PCR master mix, 1 μl primers, 5 μl DNA) 94 °C 4 min, 40× (94 °C 30 s, 55 °C 30 s, 72 °C 30 s), 72 °C 10 min [56]
Nested PCR 2 5′-CTATTGGYGGNGCYAGAGT-3′
5-GTTCATCGAAAATTCCGTGGTA-3′
381 94 °C 4 min, 40× (94 °C 30 s, 56 °C 30 s, 72 °C 30 s), 72 °C 10 min
Borrelia spp. Nested PCR 1 OspA 5′-CTTGAAGTTTTCAAAGAAGAT-3′
5′-CAACTGCTGACCCCTCTAAT-3′
487 25 µl reaction volume (12.5 μl PCR master mix, 1 μl primers, 2 μl DNA) 95 °C 5 min, 40× (95 °C 15 s, 55 °C 30 s, 72 °C 45 s), 72 °C 5 min [57]
Nested PCR 2 5′-ACAAGAGCAGACGGAACCAG-3′
5′-TTGGTGCCATTTGAGTCGTA-3′
350 95 °C 5 min, 40× (95 °C 15 s, 58 °C 30 s, 72 °C 45 s), 72 °C 5 min
Bartonella spp. Nested PCR 1 16S-23S rDNA ITS 5′-AAGTCGTAACAAGGT-3′
5′-TACTGGTKCGCTATCGGTCA-3′
1800 25 µl reaction volume (12.5 μl PCR master mix, 1 μl primers, 5 μl DNA) 95 °C 5 min, 40× (94 °C 30 s, 40 °C 45 s, 72 °C 4 min), 72 °C 10 min [58]
Nested PCR 2 5′-TTTCTCTTTCTTCAGATGATG-3
′5′-AAAGCAGGTGCTCTCCCAGAAAGCAGGTGCTCTCCCAG-3
587 25 µl reaction volume (12.5 μl PCR master mix, 1 μl primers, 2 μl DNA) 95 °C 5 min, 30× (94 °C 30 s, 60 °C 30 s, 72 °C 40 s), 72 °C 5 min