Identification of uncultured bacteria tightly associated with the intestine of the earthworm Lumbricus rubellus (Lumbricidae; Oligochaeta)

The bacteria associated with the intestine and casts of the earthworm Lumbricus rubellus Hoffmeister, 1843, were examined by direct counts, culturability studies, 16S rRNA gene clone libraries, and fluorescent in situ hybridization (FISH). A significant fraction, 24-47%, of the total numbers of prokaryotes remaining in the intestine after casting were tightly associated with the intestinal wall. Bacterial 16S rRNA gene clone libraries constructed from washed earthworm intestinal tissue suggested that the bacterial community was dominated by a few phylotypes that were either absent from, or in low abundance, in the casts. The specific phylotypes present depended on the date of sampling and included representatives of the Acidobacteria, Firmicutes, β-Proteobacteria, and one phylogenetically deep, unclassified group. Juvenile earthworms subsequently collected contained three of the four phylotypes observed in the intestine clone libraries. The Firmicutes phylotype was examined by FISH and was found to be a short rod that represented only a small fraction of the total population of the juvenile samples. These results suggested that the microbial community tightly associated with the intestine was dominated by a small number of phylotypes and that this association was opportunistic rather than obligate.