Trichogramma dendrolimi is one of the most successful biocontrol agents in China. However, an inundative condition is necessary to obtain acceptable parasitism effect. A good solution to this is the application of its thelytokous counterparts which unfortunately are scarce in field. We here report the first case of a natural T. dendrolimi population in China comprising both bisexual wasps and an extremely low proportion of thelytokous wasps. These two forms of T. dendrolimi are phylogenetically related based on the reconstructions of ITS-2 and COI genes. Also, the phylogenetic results suggested a potentially Wolbachia-drived ITS-2 variation. The expression of thelytoky was hardly affected by temperature, which might help control Asian corn borer and Dendrolimus punctatus. Wolbachia are responsible for current thelytoky according to phylogenetic analyses, antibiotic treatment and introgression experiment. We also present the third case of paternal sex ratio chromosome that restrains the expansion of Wolbachia. Moreover, the low frequency of thelytoky may be common in natural populations. Consequently if for biological control it is determined that a thelytokous strain is to be preferred, then large number of field collected females should be set up as isofemale lines, to detect the rare thelytoky.