Table 1.
Primer sets and detection probes for conventional and real-time PCRs used in the study to detect Infections caused by the parasitic species Giardia lamblia and Hymenolepis nana and those of the genera Entamoeba and Cryptosporidium
| Parasite | Target | Primer | Size, bp | Conditions | Ref. |
|---|---|---|---|---|---|
| Conventional PCR | |||||
| E. histolytica | 18S rRNA gene |
E-1 5′-GCGGTAATTCCAGCTCCA-3′ E-2 5′-GTGAAATGCTTTCGCTCTCG − 3′ |
363 | 95°C5 m [94 °C 30s 56 °C 30s 72 °C 45 s] for 26 × 72 °C 6 m | This study |
| E. dispar | 264 | ||||
| E. coli | 435 | ||||
| E. nana | 645 | ||||
| E. muris | 453 | ||||
| G. lamblia | tpi gene (1) | tpi(A-for) 5′-GGAGACCGACGAGCAAAGC-3′ tpi (A-rev), 5′-CTTGCCAAGCGCCTCAA-3′ (Assemblage A)tpi(B-for), 5′-AATAGCAGCACARAACGTGTATCTG-3’tpi(B-rev), 5′-CCCATGTCCAGCAGCATCT-3′(Assemblage B) |
184 81 |
95°C5 m [94 °C 30s 62 °C 30s 72 °C 30s] for 38 × 72 °C 7 m | [26] |
|
H. nana H. dimenuta |
18S rDNA gene, ITS1, | F3 (5′-GCGGAAGGGATACTTACACGTTC-3′) R3 (5-'GCTCGACTCTTCATCGATCCACG-3′) |
646 754 |
95°C5 m [95 °C 20s 60 °C 30s 72 °C 45 s] for 35 × 72 °C 7 m | [27] |
| Cryptosporidium spp | COWP gene(2) | cry15: 5′-GTAGATAATGGAAGAGATTGTG-3 cry9: 5′-GGACTGAAATACAGGCATTATCTTG-3’ | 553 | 95°C5 m [94 °C 30s 60 °C 30s 72 °C 30s] for 38 × 72 °C 7 m | [28, 29] |
| Real-time PCR | |||||
| C. parvum spp |
C. parvum-specific 452-bp DNA fragment. |
Cr-F 5′-CGCTTCTCTAGCCTTTCATGA-3′ Cr-R 5′-CTTCACGTGTGTTTGCCAAT-3′ CryptoTP1 Tex Red- 5′-CCAATCACAGAATCATCAGAATCGACTGGTATC-3′ BHQ2 | 138 | 95°C1 5s [95 °C 15 s 60 °C 30s 72 °C 30s] for 40x | [25] |
| G. lamblia | SSU RNA gene (3) | Giardia 80F 5′-GACGGCTCAGGACAACGGTT-3′ Giardia 127R 5′-TTGCCAGCGGTGTCCG-3′ Giardia-105 T FAM-5′-CCCGCGGCGGTCCCTGCTAG-3′-BHQ 1 | 62 | 95 °C 15 s [95 °C 15 s 60 °C 30s 72 °C 30s] for 40x | [25] |
(1) tpi: triosephosphateisomerase, (2) COWP: Cryptosporidium oocyst wall protein, (3) SSU RNA: Small subunit RNA.