Age-dependent changes in intestinal microflora of honeybee
Remi Kasahara1, Jun Nakamura2, Yoshikazu Koizumi3, Ayako Mitsui3, and Masami Sasaki4. (1) Graduate School of Agriculture, Tamagawa University, Machida, Tokyo, 1948610, Japan, (2) Honeybee Science Research Center, Tamagawa University, Machida, Tokyo, Japan, (3) Environmental Engineering Center Co., Ltd., Machida, Tokyo, 1948610, Japan, (4) Laboratory of Entomology, Faculty of Agriculture, Tamagawa University, Machida, Tokyo, 1948610, Japan
The intestinal microflora of honeybee was investigated by means of the PCR-DGGE (denaturing gradient gel electrophoresis) method based on the sequence-specific separation of PCR-derived rRNA gene amplicon, which have proven useful in analyses of wide ranged studies in microbial ecology. Entire intestinal contents of adult honeybees were removed with gut wall under sterile conditions and the whole genomic DNA was isolated. PCR was used to amplify 16S rRNA genes from the DNA with a set of bacterial specific GC-338F and universal primers. The former one contains a 40 base GC-rich sequence at the 5’-end. The result of DGGE profiles and the DNA sequence analyses confirmed that the intestinal microflora had already existed in the newly emerged workers (day 0), however, it was very simple at day 0 and consisted of only one or two common bacteria. The DNA sequence of one of those showed the homology to Lactobacillus alvei strain 1G2 with 97% similarity. Then the microflora tended to be complex with age, and in the foragers, the composition of bacteria was varied besides the several common ones. The age dependent changes in higher diversity of the intestinal microflora in foragers are probably due to the higher accessibility to the sources of bacteria, namely foods, nestmates, combs, and outside food sources. We discuss the effects of nutritional status and trophallaxis with other nestmates on the individual intestinal microflora, as well as the effects of season, location and food sources on the colony level intestinal microflora.