In this article, the composition and distribution of bacteria associated with the gut of Tenebrio molitor (L.) larvae were investigated using both culture-dependent and culture-independent denaturing gradient gel electrophoresis (DGGE) methods. This work compares bacterial species associated with four different parts of T. molitor larvae gut: foregut, anterior midgut, posterior midgut, and hindgut. Five genera, Weissella, Lactococcus, Rahnella, Cronobacter, and Enterococcus, were isolated using nutrient agar. All of these strains were present in the posterior midgut and hindgut. The strains with milk-clotting activity in selective casein-plates assay were sequenced and identified as species of genera Weissella and Lactococcus, and those with proteolytic activity as Rahnella and Cronobacter, implying that they may be involved in protein utilization. But none of these strains showed cellulolytic activity. In DGGE experiment, 19 isolated bands belonging to nine taxa (Spiroplasma, Lactococcus, Lactobacillus, Bacillus, an uncultured Bacillaceae, Clostridium, Enterobacter, Pantoea, and an uncultured Clostridium) were extracted and identified from DGGE gels. These species could be assigned to three phyla Tenericutes, Firmicutes, and Proteobacteria. According to the DGGE analysis, the bacterial communities of the four gut regions exhibited some differences, with the hindgut showing the highest bands abundance and diversity. The genus Spiroplasma, which is generally regarded as pathogen or male-killing bacteria in insects, had a high abundance in the gut environment, their potential role is worthy of a further study.