While the G2 PhyloChip is an excellent tool for identifying known bacteria, it contains only 300 archaeal sequences, which were not utilized because bacterial-specific primers were used. Furthermore, there is currently no microarray that is designed to identify protozoa or fungi. Next generation (high-throughput) sequencing is needed to validate the bacterial population findings of the present study, as well

as identify the protozoal, archaeal and fungal populations present in the moose rumen. The PhyloChip, like all methods that do not rely on culturing, cannot be used to differentiate between transient and colonizing species. It can be assumed that some species found in the moose are simply passing through the digestive tract, having been picked up from the environment, and are not colonizing the tract. Despite

this, GSK126 these transient CB-839 research buy bacteria may still have an impact on the dynamics within the rumen, and it is important to take a holistic approach when looking at mixed environmental samples. It is also possible that some of these unclassified bacteria which are presumed transient, such as the soil or water clones, are actually colonizing the moose digestive tract and are simply unique to moose. Methods Sample collection All samples were obtained with permission of licensed hunters through the Vermont Department of Fish and Wildlife. Whole rumen (R) and colon (C) contents were collected from moose shot during the October 2010 moose hunting season in Vermont. Samples were collected by hunters within 2 h, if not sooner,

of death and put on ice immediately. Hunters were given a written set of instructions about sample collection, and had been instructed verbally as well, to fill the collection containers with material taken Tolmetin from well inside the rumen and colon, and to seal the container quickly to minimize overexposure to oxygen. Samples were then transferred to the laboratory within 24 h, and stored at −20°C until DNA extraction. A total of eight rumen and six colon samples (Table 3) were collected from eight moose. Twelve of the samples were paired rumen and colon contents from the same animal, and two rumen samples did not have corresponding colon samples. Moose were weighed and aged, by examining the wear and replacement of the preLB-100 cell line molars and molars of the lower jar, by Vermont Fish and Wildlife biologists at the mandatory reporting stations. Table 3 Statistics for samples taken from moose shot in October 2010 in Vermont during the moose hunting season Moose Sample location Sample name Gender Weight, dressed carcass (kg) Approx. age (yr) 1 Rumen 1R F 185 1   Colon 1C       2 Rumen 2R F 244.55 3   Colon 2C       3 Rumen 3R M 186.36 2   Colon 3C       4 Rumen 4R M N/A N/A 5 Rumen 5R M 319.09 4 6 Rumen 6R F 259.55 3   Colon 6C       7 Rumen 7R M 301.36 4   Colon 7C       8 Rumen 8R M 405.