Helicobacter pylori and other Helicobacter species DNA in human bile samples from patients with various hepato-biliary diseases

Table 1 List of primers used for the study

Primer	Sequence(5’→3’)	Productsize
		(bp)
Heli-nestS	5’ATTAGTGGCGCACGGGTGAGTAA3’	1 300
Heli-nestR	5’TTTAGCATCCCGACTTAAGGC3’
Heli-S	5’GAACCTTACCTAGGCTTGACATTG3’	480
Heli-R	5’GGTGAGTACAAGACCCGGGAA3’
16S-rRNA.F	5’TAAGAGATCAGCCTATATGTCC3’	534
16S-rRNA.R	5’TCCCACGCTTTAAGCGCAAT3’
UreA-S	5’GCCAATGGTAAATTAGTT3’	411
UreA-R	5’CTCCTTAATTGTTTTTAC3’
CagA-S	5’CCATGAATTTTTGATCCGTTCGG3’	349
CagA-R	5’GATAACAGGCAAGCTTTTGAGAGGGA3’

Table 2 Conditions of polymerase chain reactions used in the study

Target gene	Initial denaturation step	Temperature o denaturation, annealing and	Cycle	Final extension Step extension
Helicobacter	94 °C for 5 min	94°C for 30 s
Spp,16S rRNA		55°C for 30 s	35	72°C for 7 min
				72°C for 1.5 min
Second	94°C for 5 min	94°C for 30 s
amplification		60°C for 30 s	35	72°C for 7 min
step		72°C for 30 s
Helicobacter	95°C f	94°C for 30 s
pylori, 16S rRNA		56°C for 30 s	40	72°C for 5 min
				2°C for 1 min
ureA	95°C for 5 min	94°C for 30 s	35	72°C for 6 min
		52°C for 30 s		72°C for 1 min
cagA	95°C for 5 min	94°C for 1 min
		52°C for 1 min	35	72°C for 10 min
		72°C for 2 min

Table 3 Details of Helicobacter genus, H pylori 16S rRNA, ureA, cagA positivity in the study subjects

No	Category (n=60)	Helicobacter Genus positivity n(%)	Hpylori16S rRNA positive n(%)	Ure A positive n(%)	Cag A positive n(%)
1	Group I	29(96.7)	10(33.3)	10(33.3)	09(30)
2	Group II	02(6.6)	01(3.3)	01(3.3)	01(3.3)