Objective To explore the relationship between virulence genes and antibiotic resistance in Helicobacter pylori (H. pylori) isolates in children. Methods The stains of H. pylori were isolated from antral mucosa in 90 pediatric patients with upper gastrointestinal symptoms. E-test and K-B test were used to detect the antibiotic resistance of H. pylori isolates. PCR was performed to detect the cagA, vacA and iceA genes of H. pylori. Results Among these 90 strains of H. pylori isolated from antral mucosa, there were 8 strains (8.9%) resistant to clarithromycin, 31 strains (34.4%) resistant to metronidazole, 12 trains (13.3%) resistant to clarithromycin and metronidazole, and 39 strains (43.3%) not resistant to antibiotics. There were no strains  resistant to amoxicillin or furazolidone. The positive detection rate of cagA gene, vacAs1a, vacAs1c, vacAm1, and vacAm2 genes were 93.3% (84/90), 77.8% (70/90), 22.2% (20/90), 32.2% (29/90), and 67.8% (61/90), respectively. The positive detection rates of vacAs1a/m1, vacAs1a/m2, vacAs1c/m1, and vacAs1c/m2 genes were 30.0% (27/90), 51.1% (46/90), 3.3% (3/90), and 16.7% (15/90), respectively. The positive detection rates of iceA1 and iceA2 genes were 87.7% (79/90) and 7.8% (7/90), respectively. There was no difference in positive detection rate of H. pylori virulence genes among clarithromycin resistance group, metronidazole resistant group, clarithromycin and metronidazole double resistance group and antibiotics sensitive group (P > 0.05). Conclusion There was no relationship between virulence gene and antibiotic resistance in H. pylori clinical isolates in children.