Affiliation:
1. Laboratory of Rumen Microbiology, STAFF-Institute, Tsukuba, Ibaraki 305-0854,1 and
2. Department of Animal Sciences, University of Illinois at Urbana-Champaign, Urbana, Illinois 618012
3. Japan Collection of Microorganisms, The Institute of Physical and Chemical Research, Wako, Saitama 351-0198,3 Japan, and
Abstract
ABSTRACT
A set of PCR primers was designed and validated for specific detection and quantification of
Prevotella ruminicola
,
Prevotella albensis
,
Prevotella bryantii
,
Fibrobacter succinogenes
,
Selenomonas ruminantium-Mitsuokella multiacida
,
Streptococcus bovis
,
Ruminococcus flavefaciens
,
Ruminobacter amylophilus
,
Eubacterium ruminantium
,
Treponema bryantii
,
Succinivibrio dextrinosolvens
, and
Anaerovibrio lipolytica
. By using these primers and the real-time PCR technique, the corresponding species in the rumens of cows for which the diet was switched from hay to grain were quantitatively monitored. The dynamics of two fibrolytic bacteria,
F. succinogenes
and
R. flavefaciens
, were in agreement with those of earlier, culture-based experiments. The quantity of
F. succinogenes
DNA
,
predominant in animals on the hay diet, fell 20-fold on the third day of the switch to a grain diet and further declined on day 28, with a 57-fold reduction in DNA. The
R. flavefaciens
DNA concentration on day 3 declined to approximately 10% of its initial value in animals on the hay diet and remained at this level on day 28. During the transition period (day 3), the quantities of two ruminal prevotella DNAs increased considerably: that of
P. ruminicola
increased 7-fold and that of
P. bryantii
increased 263-fold. On day 28, the quantity of
P. ruminicola
DNA decreased 3-fold, while
P. bryantii
DNA was still elevated 10-fold in comparison with the level found in animals on the initial hay diet. The DNA specific for another xylanolytic bacterium,
E. ruminantium,
dropped 14-fold during the diet switch and was maintained at this level on day 28. The concentration of a rumen spirochete,
T. bryantii,
decreased less profoundly and stabilized with a sevenfold decline by day 28. The variations in
A. lipolytica
DNA were not statistically significant. After an initial slight increase in
S. dextrinosolvens
DNA on day 3, this DNA was not detected at the end of the experiment.
S. bovis
DNA displayed a 67-fold increase during the transition period on day 3. However, on day 28, it actually declined in comparison with the level in animals on the hay ration. The amount of
S. ruminantium
-
M. multiacida
DNA also increased eightfold following the diet switch, but stabilized with only a twofold increase on day 28. The real-time PCR technique also uncovered differential amplification of rumen bacterial templates with the set of universal bacterial primers. This observation may explain why some predominant rumen bacteria have not been detected in PCR-generated 16S ribosomal DNA libraries.
Publisher
American Society for Microbiology
Subject
Ecology,Applied Microbiology and Biotechnology,Food Science,Biotechnology