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Using specific primers, we measured the infection status of Wolbachia, WO, Cardinium, and Spiroplasma in Chilo suppressalis (Walker) (Lepidoptera: Crambidae) collected from 14 locations in China, and conducted phylogenetic analysis of the three endosymbionts. The results showed that eight geographical populations of C. suppressalis were infected by Wolbachia. The infection rates of Wolbachia in four geographical populations had no significant difference between 2011 and 2012 (P>0.05). There were five geographical populations infected with Cardinium and seven geographical populations infected with Spiroplasma. Seven populations of C. suppressalis carrying Wolbachia were also infected with WO. The maximum parsimony tree shows that Wolbachia from C. suppressalis, Pieris rapae, Coliaserate poliographus and Tetranychus urticae fall into the same clade. Cardinium from C. suppressalis joins together with Cardinium from Oppiella nova. Spiroplasma in C. suppressalis has a close relationship with that of Fannia manicata. In conclusion, infections of three endosymbionts showed regional differences, and there were eight geographical populations infected with more than one endosymbiont. Our study found, for the first time, double infection with Wolbachia and Spiroplasma in one sample from Xinzhou. Phylogenetic analysis of the three endosymbionts proved that the endosymbionts can transfer horizontally between different individuals of the same or different hosts. [ABSTRACT FROM AUTHOR]