Table 1.
Primers and PCR cycle conditions used for the molecular characterization of pathogens and hippoboscid flies (Diptera: Hippoboscidae) collected from domestic and wild ruminants in Austria
| Organism | Target gene | Primers | Sequence (5'–3') | Length (bp) | PCR cycle conditions | References |
|---|---|---|---|---|---|---|
| Anaplasmataceae | 16S rRNA | EHR16SD_for | GGT ACC YAC AGA AGA AGT CC | 345 | 95 °C/2 min; 35 cycles: 94 °C/1 min, 54 °C/30 s, 72 °C/30 s; 72 °C/5 min | [40] |
| EHR16SR_rev | TAG CAC TCA TCG TTT ACA GC | |||||
| Borrelia | 16S rRNA | Borr_allg_for | ACG CTG GCA GTG CGT CTT AA | 674 | 94 °C/2 min; 6 cycles: 94 °C/1 min, 45 °C/1.5 min, 72 °C/75 s; 36 cycles: 94 °C/1 min, 51 °C/1.5 min, 72 °C/75 s); 72 °C/5 min | [39] |
| Borr_allg_rev | CTG ATA TCA ACA GAT TCC ACC C | |||||
| Bartonella | gltA | BhCs.781p | GGG GAC CAG CTC ATG GTG G | 379 | 94 °C/5 min; 40 cycles: 94 °C/1 min, 54 °C/1 min, 72 °C/1 min; 72 °C/10 min | [38] |
| BhCs.1137n | AAT GCA AAA AGA ACA GTA AAC A | |||||
| Filarioidea | COI | COlint-F | TGA TTG GTG GTT TTG GTA A | 668 | 94 °C/2 min; 8 cycles: 94 °C/45 s, 51 °C/45 s, 72 °C/1.5 min; 25 cycles: 94 °C/45 s, 45 °C/45 s, 72 °C/1.5 min; 72 °C/7 min | [42] |
| COlint-R | ATA AGT ACG AGT ATC AAT ATC | |||||
| Trypanosomatida | 18S rRNA | Tryp_18S_F1 (Nest 1) | GTGGACTGCCATGGCGTTGA | 960 | 94 °C/5 min; 35 cycles: 94 °C/1 min, 56 °C/1 min, 72 °C/1 min; 72 °C/5 min (Nest 1) | This study |
| Tryp_18S_R1 (Nest 1) | CAGCTTGGATCTCGTCCGTTGA | |||||
| Tryp_18S_F2 (Nest 2) | CGATGAGGCAGCGAAAAGAAATAGAG | 94 °C/5 min; 25 cycles: 94 °C/1 min, 56 °C/1 min, 72 °C/1 min; 72 °C/5 min (Nest 2) | ||||
| Tryp_18S_R2 (Nest 2) | GACTGTAACCTCAAAGCTTTCGCG | |||||
| Piroplasmida | 18S rRNA | BTH-1F (Nest 1) | CCT GAG AAA CGG CTA CCA CAT CT | 561 | 94 °C/2 min; 40 cycles: 95 °C/30 s, 68 °C/1 min, 72 °C/1 min; 72 °C/10 min (Nest 1) | [41] |
| BTH-1R (Nest 1) | TTG CGA CCA TAC TCC CCC CA | |||||
| G-2_for (Nest 2) | GTC TTG TAA TTG GAA TGA TGG | 94 °C/2 min; 40 cycles: 95 °C/30 s, 60 °C/1 min, 72 °C/1 min; 72 °C/10 min (Nest 2) | ||||
| G-2_rev (Nest 2) | CCA AAG ACT TTG ATT TCT CTC | |||||
| Lepidoptera | COI | LepF1 | ATT CAA CCA ATC ATA AA | 648 | 94 °C/2 min; 6 cycles: 94 °C/1 min, 45 °C/1.5 min, 72 °C/75 s; 36 cycles: 94 °C/1 min, 51 °C/1.5 min, 72 °C/75 s); 72 °C/5 min | [37] |
| LepR1 | TAA ACT TCT GGA TGT CAA AAA |