Terminal Restriction Fragment Length Polymorphism (T-RFLP) and Denaturant Gradient Gel Electrophoresis (DGGE) have been used to describe variations and diversity of the microbiota in the intestinal tract in broilers [8–10]. However, when it comes elucidate the phylogenetic diversity in
the intestinal microbiota at species level, these methods are not sensitive and specific enough. By traditional culture methods only culturable genera are detected, and these are estimated to be about 1% of all genera present in the microbiota , whereas DGGE only detects species that represent more than 1% of the total microbiota Wortmannin , and in T-RFLP, sequence redundancy at the cleaving side may generate fragments of the same
length from various species. A more comprehensive description of the distribution of species in the microbiota can be done by Sanger sequencing of 16S rDNA libraries. With this method individual species are arranged into Operational Taxonomic Units (OTU) based on > 98% similarity of 16S rDNA sequences [8, 13], but as these methods are very laborious, only the most dominating species are detected. A much deeper investigation of the microbiota has been achieved with the introduction of second generation sequencing technology, such as 454 pyrosequencing, selleckchem where massive parallel sequencing of short hyper variable regions within the 16S rDNA is performed [14–16]. Using this technology, a 16S rDNA selleck chemicals llc library may be sequenced in one run; generating a large number of sequence reads that allows a much deeper insight in the distribution of species. Although the generated sequences do not cover the whole gene, Huse et al.  GNAT2 were able to achieve a 99% correlation of identification, when compared with full
length sequencing of a library from the human microbiota. The microbiota of laying hens experiencing nutritional stress has been investigated by 454 pyrosequencing . In this study, the authors described the changes in the microbiota induced by different molting methods, where hens were given different feed or being starved. By starving the layers, they observed a decrease in species diversity of the caecal microbiota which was not found in hens receiving a diet with high fiber content. With the change to more welfare friendly cage systems, laying hens are now going to be housed in larger groups of 60 birds, rather than 4-6 birds as seen in conventional battery cages. Whether these changes in group size, increased contact between individuals or change in behavior may also have influence on the diversity of the species in the intestinal tract or in the oviduct, have not been investigated.