A duplex tetra-primer ARMS-PCR assay to discriminate three species of the Schistosoma haematobium group: Schistosoma curassoni, S. bovis, S. haematobium and their hybrids

Table 3 Tetra-primer amplification refractory mutation system-PCR primer sets for the identification of S. haematobium, S. bovis and S. curassoni and their hybrids

DNA target	T-ARMS-PCR primer set^a	Sequence	Working concentration (µM)	Melting temperature (Tm; °C)	Length (bp)
ITS
Outer	OR-ITS	TCGTGCGTATTACACACACCATCGGTACAAACC	12	64	33
Outer	OF-ITS	GCATGCAAATCCGCCCCGTTATTGTTCCT	12	64.7	29
Inner	IR-Sh-ITS	TGTGGCCGGACTATTAGGACGGAGCCGTC	15	67.5	29
Inner	IF-Sb/Sc-ITS	CGCATATCAACGCGGGTTGCTGGGCA	15	67.3	26
18S
Outer	OR-18S	GCACCAGACTTGCCCTCCAATTGGTCC	12	65.2	27
Outer	OF-18S	GCATTTATTAGAACAGAACCAAYCGGGCG	12	59.8/61.5	29
Inner	IR-Sc/Sh-18S	ATTTGAAAGATCCGTCGCCGACAAAA	15	59.3	26
Inner	IF-Sb-18S	TGGATAACTTTACTGATCGCAGTCGGACC	15	61.2	29

OF and OR define forward and reverse external primers, respectively. IF and IR define internal forward and reverse primers, respectively. The outer primers for each target generate the internal control Schistosoma sp. amplicon, whereas the inner primer together with the outer primers generate the species-specific amplicons
^aIR-Sh-ITS + OF-ITS generates the ITS S. haematobium amplicon. IF-Sb/Sc-ITS + OR-ITS generates the ITS S. bovis or S. curassoni amplicon. IR-Sc/Sh-18S + OF-18S generates the 18S S. haematobium or S. curassoni amplicon. IF-Sb-18S + OR-18S generates the 18S S. bovis amplicon

ISSN: 1756-3305