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1440
THE AMERICAN JOURNAL OF CLINICAL NUTRITIoN
Vol. 23, No. 11, November, 1970, pp. 1440-1450
Printed in U.S.A.
Normal Flora Rumen Bacteria1
M. P. BRYANT,2 PH.D.
LL HERBIVOROUS ANIMALS have an ex-
panded part of the alimentary tract
where bulky fibrous foods, which are rich
in cellulose and similar polysaccharides,
can be delayed in passage in order to
undergo the extensive microbial fermenta-
tion that is necessary for utilization. In
ruminants the expanded part of the tract
is represented by the rumen. Ingested food
passes directly from the esophagus to the
reticulorumen, which represents about 85%
of the total stomach capacity and contains
digesta equal to about 10-20% of the
animal’s weight. The food stays in the
rumen until the microbial fermentation
has brought about the digestion of about
70-85% of the total digestible dry matter.
The undigested food residues and microbial
cells produced in the fermentation con-
tinuously pass out of the reticulum and
into the omasum and, hence, to the
abomasum, or true stomach and intestine,
where digestion of microbes and feed
residues proceeds much as it does in
nonruminant animals.
The fermentation represents a very com-
plex symbiotic association involving the
animal and specific groups of microorga-
nisms that have evolved with the animal.
The chief groups of microorganisms in-
clude certain ciliate protozoa, which are
not found elsewhere in nature, some
flagellates, and a very complex mixture of
bacteria consisting mainly of nonspore-
forming anaerobes. Although the protozoa
contribute significantly to the fermentation,
they are not essential. My discussion will
1 From the Departments of Dairy Science and
Microbiology, University of Illinois, Urbana, Illinois
61801.2 Professor of Microbiology.
mainly concern the small bacteria that
always account for much of the activity.
Most of the information has been obtained
from cattle and sheep. Other ruminants
appear to support a similar flora but
relatively little information on these has
been obtained. A number of comprehen-
sive reviews on various aspects of rumen
microbiology are available. (See (1, 2, and
3) for further references.) Thus, no attempt
will be made to present complete references
in this paper.
The Rumen Environment
The rumen can be likened to a highly
efficient continuous culture system for
growth of anaerobic microorganisms. Food
and water supply, temperature and osmotic
pressure, mixing, and outflow of undigested
residues and microorganisms are relatively
constant. The pH is held relatively con-
stant, usually 6-7, by the buffering action
of a large amount of secreted saliva, which
is high in sodium and potassium bicarbo-
nate and urea, by absorption through the
rumen wall into the blood stream of acids,
and by ammonia produced in the fermen-
tation. The oxidation-reduction potential
is maintained at about -400 my because
of the intensive microbial activity and the
entrance of only a small amount of
oxygen. Gas production is rapid and con-
sists of about 50-70% of CO2 and the rest
is mainly methane. Because of the very
rapid fermentation of soluble proteins and
carbohydrates of the feed, carbohydrate
energy sources for microbes and protein
are mainly present in the particulate frac-
tion of the digesta during the greater part
of time between feedings. Soluble materials
in rumen fluid include a large amount of
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Normal Flora-Rumen Bacteria 1441
ammonia available as a nitrogen source for
bacteria and the volatile acids and some
other organic acids available as carbon
sources, but not usually available as energy
sources, for the bacteria. Minerals and
vitamins are also present in the fluids.
One of the major ways in which the
rumen differs from a continuous culture
apparatus is that the ingesta is quite
stratified (4). Thus, much more of the
fibrous materials and digestible dry matter
are present in dorsal contents than in
ventral contents. Also, the turnover time
of particles such as hay is longer, i.e.,
about 1-2 days, as compared with 0.3-0.8
day for materials in the fluids (1).
Based on turnover times one can see that
the bacteria in the rumen, even if associ-
ated solely with the more rapidly passing
liquid phase, would only have to grow
with a very slow mean generation time,
about 7 hr, to be maintained. However,
about twice as many bacteria per unit
weight are present in the dorsal contents,
which contains most of the undigested
fibrous particulate materials, rather than
the more liquid ventral material (4). Thus,
the mean generation time needed to main-
tain bacteria in the rumen is probably
much longer than 7 hr even under condi-
tions when rates of passage are very rapid.
Although maximum growth rates of few
rumen bacteria have been determined, it
seems certain that they almost always grow
at a much slower rate than is their poten-
tial rate.
General Functions of the Bacteria
A typical balance for the fermentation
of carbohydrate in the rumen as calculated
by Wolin (5) is given in the following
equation:
57.5(C,H120,) -* 65 acetate + 20 propionate
+ 15 butyratc + 60 CO2 + 35 CH4 + 25 H20
Small amounts of n-valerate and, sometimes,
n-caproate are also produced. Energy con-
version allows microbial growth (dry
weight) equal to about 10-20% of the
carbohydrate fermented (1, 6). The volatile
acids produced are the main source of
energy for the ruminant whereas the
methane, containing about 8% of the gross
energy of the animal’s diet, is lost.
Protein synthesis by rumen bacteria is
very important to the animal (7). The more
soluble proteins of the diet are very rapidly
hydrolyzed to peptides and amino acids
and either converted to microbial protein
or further catabolized to ammonia, volatile
and other organic acids (i.e., the aromatic
acids produced from aromatic amino acids),
carbon dioxide, and sulfide. Practically all
of the bacteria can utilize ammonia as the
main source of nitrogen, and those bacteria
requiring amino acids require only one or
a few. If too little energy is available for
efficient microbial growth, much of the
ammonia is absorbed into the blood
stream, converted to urea, and excreted.
Thus, a proper balance of carbohydrate
energy source and nitrogen is necessary in
the diet in order for the animal to utilize
nitrogen efficiently and to obtain a proper
supply of essential amino acids.
A significant amount of urea from the
blood and saliva is returned to the rumen
and a very active microbial urease converts
it to ammonia and carbon dioxide.
Because under usual conditions most of
the digestible dietary protein is converted
to microbial protein in the rumen, the
distribution of essential amino acids in the
diet of ruminants is of relatively little con-
sequence as compared to nonruminant
animals. In fact, due to the ability of the
bacteria to synthesize amino acids using
ammonia nitrogen, the ruminant has no
requirement for amino acids, and most of
the nitrogen in the diet can be supplied as
urea or other nonprotein nitrogen com-
pounds.
It is of interest that a significant number
of the rumen bacterial species cannot
efficiently utilize amino acids or peptides
but require ammonia as the nitrogen
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1442 Bryant
source (8, 9), whereas some others utilize
peptides or ammonia but do not effectively
utilize free amino acids (10).
Rumen bacteria also are important in
ruminant fat metabolism. Glycerides and
other lipids such as the galactosyl glyc-
eiides of clover lipids are rapidly hydro-
lyzed and the glycerol or galactose is
fermented, the former mainly to propionic
acid (11). The unsaturated fatty acids are
largely hydrogenated but fatty acids are
not degraded by rumen bacteria. Rumen
bacteria contain relatively large amounts of
odd-numbered carbon straight-chain and
branched-chain saturated fatty acids (11)
and these are biosynthesized by some
species from carbohydrate or amino acid
carbon, but a number of species require
straight-chain volatile acids such as n-
vales-ate or branched-chain acids such as
isovalera te, isobutyrate, or 2-methylbuty-
rate, or both, in order to grow and bio-
synthesize the longer chained acids (12).
Relatively large amounts of these acids are
presellt in ruminant fat (11).The ruminant animal is not dependent
on an exogenous supply of B-vitamins or
vitamin K as they are all biosynthesized by
runien bacteria (13).
Culture and Nutrition of the Bacteria
Dus-ing the first half of the century many
workers attempted the pure culture of
bacteria, functional in the rumen, with
maj 01� emphasis on cellulolytic bacteria;
however, it was not until 1946 that Hun-
gate (14) had unqualified success in ob-
taining pure cultures of the important
cellulolytic bacteria, Bacteroides succino-
genes and members of the genus Rum mo-
coccus. The success of the methods was due
to the development of the roll-tube
technique, which allows very good anaero-
bic conditions to be continuously main-
tamed, and to the use of a prereduced
medium with a composition similar to that
of the natural environment of the orga-
nisms. Although Hungate was, at first,
mainly interested in cellulolytic bacteria
and added finely dispersed cellulose as
the main energy source in the media, he
found that very large numbers of noncellu-
lolytic bacteria grew. This was especially
true when a sugar such as glucose was
added.
It is surprising that little definitive work
has been published on the agar media best
suited for the relatively nonselective erni-
meration and isolation of bacteria in the
various anaerobic habitats in nature and!,
also, that anaerobic techniques often have
been inadequate. However, after working
in Hungate’s laboratory during the time
cellulolytic anaerobes and treponemes
(Borrelia sp.) were routinely isolated!
(14, 15), we modified slightly the anaerobic
techniques and developed the RGCA
medium (16) and, latem-, medium 98-5 that
appears to be as good as any medium for
isolation and enumeration of rumen bac-
teria (17). The latter medium is only
slightly modified from that of Hungate
(14), and similar media have been indi-
cated to be the best available for intestinal
and fecal anaerobes of man and various
animals (18), total anaerobic counts of
anaerobic sewage digestors (19), and the
most sensitive anaerobes known, i.e., the
methanogenic bacteria (20).
Medium 98-5 was developed on the bases
of a large number of experiments in which
various additions, dieletions, and levels of
ingredients were te,ted via colony counts
of rumen contents and isolation and pre-
sumptive identification of large numbers of
strains of rumen bacteria (17). The medium
contains small amounts of glucose, cello-
biose, and starch as addled! energy sources.
The three are necessary because some
carbohydrate-fermenting an aerobes, as i ndi-
cated later, utilize only one of these.
Addition of other energy souices such as
lactate or glycerol would be necessary for
growth of a few species. Energy sous-ces
should be added at a low level to keep gas
production low and! colonies and spreading
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Normal Flora-Rumen Bacteria 1443
F,, = observed difference in electromotive force
between electrode and normal hydrogen electrode.
small so that accurate counts and effective
isolation can be made from tubes contain-
ing many colonies or incubated for long
times. Clarified rumen fluid is addled as a
source of many growth factors including
some that are not found in the usual
bacteriological media. The main buffer in
the medium (pH about 6.7) is bicarbonate-
carbonic acid. This is provided by the
addition of NaHCO3 and maintenance of
a CO2 or 1: 1 C02-H2 gas phase. A carbon
dioxidle gas phase of 50% or more is
1-ecommended because many of the rumen
anaerobes, and especially those that pro-
duce succinate as a major product, require
substrate levels for growth (16, 21). A low
oxidation-redluction potential is obtained
by adding a combination of cysteine and
sulfide as reducing agents. In our hands,
inclusion of 0.025% of Na2S�9H2O in addi-
tion to cysteinc approximately doubles
colony counts (17). Resazurin, 0.0001%, is
usually added as a crude indicator of
anaerobiosis. If reduced to the colorless
state, it indicates an Eh (see footnote 3)
more negative than about -75 my. Actis-
ally, the medium rapidly attains an E,, of
about -250 my or lower, i.e., the Eh at
which phenosafranine is partially ieduced
and indligo disulfonate is completely re-
ducedl (2).
Many of the rumen anaerobes were
shown to require factors present in rumen
fluid i)ut not found in sufficient amounts
in ingredients commonly added to bacteri-
ological media or in feeds consumed by
ruminants. Work done mainly at Beltsville
in the 1950’s and early 1960’s established
the nature and functions of most of these
growth factors and showed that the
greater number of strains of most species
of rumen anaerohes could be grown in
defined media (2, 3, 9, 12, 21). An exception
is the methanogenic bacterium that requires
an as yet unidentified growth factor (22).
The work suggested that rumen fluid in the
medium for enumeration and isolation of
most bacteria could be replaced by more
readily available and better standardized
ingredients; and this was subsequently
shown to be possible (23).
Medium 10 was essentially identical with
the rumen fluid medium (17) except that
the rumen fluid was replaced with a number
of ingredients (23). Small amounts of
trypticase and yeast extract were adided as
sources of vitamins, peptides, and! amino
acids. For most species, these could prob-
ably be replaced by B-vitamins alone, or
B-vitamins plus one or a few amino acids
(9). Very few rumen bacteria i-equire
nucleic acid degradation products. Hemin
was added! because it is essential foi growth
of most strains of Bacteroides ruminicola,
which is usually very numerous (24). The
volatile fatty acids, acetate, n-valerate,
isovalerate, 2-methylbutyrate, and isobuty-
i-ate were added as one or more of these
are either essential or highly stimulator) to
growth of many species of rumen anaei-obes
(2, 9, 22). Vitamin K, or menad!ione, is
essential to growth of some strains of
Bacteroides melaninogenicus (25) but was
not addled to the medium as this species
is a minor component of the rumen floia.
The establishment of the requirement of
volatile fatty acids and! heme for growth
of many rumen bacteria allowed recogni-
tion of important mici-obial interactions in
that these factors are produced by other
species of rumen bacteria. For example, the
branched!-chain volatile acids required by
many species are prod!uced by oxidative
decarboxylation of the corresponding
branched-chain amino acids by other spe-
cies (3).
Some Species of Rumen Bacteria
The rumen contains a very large number
of species (1). Many aeiobic and faculta-
tively anaerobic species are present in small
numbers and! are not metabolically active
un(lel riimen conditions, or are present in
too small numbers to be functionally
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1444 Bryant
significant. For example, coliforms are
usually present in numbers of about 1,000-
10,000/g and Streptococcus bovis, often the
most numerous facultative anaerobe, is
usually present in numbers of l05-lOT/g.
Many of the more important species, the
nonsporeforming anaerobes, are usually
present in numbers of 108-1010/g in very
young animals (26) as well as in adults (2).
Under usual conditions, no single species
dominates the flora. For example, among
about 50 randomly, pure cultured strains
from well isolated colonies from high
dilutions of rumen contents, at least 10-
15 quite different species representing
about 6-10 genera are commonly ob-
tained. Under a few conditions such as
certain regimes with very high grain feed-
ing a few species may dominate (see (23)).
Many of the dominant species utilize
TABLE I
Some features of more numerous rumen
anaerobic bacteria-gram-negative, non-
motile rods of the genus Bacteroides
F eature B.ruminicola
B.succinogenes
B.amylop/sUus
Energy sources
Glucose + +
Maltose-starch + ZF +Cellulose - +Xylan
Others
Fermentation
products”
Cytochrome b
NH3 from amino
+ManyS,A,F,P(-C02)
+
+
-
FewS,A,F(-C02)
+-
-
NoneS,A,F
(-C02)
-
-
acids
Some nutrient re-
quirements
Heme required
Acids requiredb
±
Stimula-
lation
-
+-
-
NH3 required
Animal diet where
numerous
-
Many
+Many
+High
grain
= succinate, A = acetate, F = formate,
P = propionate, (-CO2) = CO2 uptake. b
Valerate or longer straight-chain acid and/or
isobutyrate or 2-methylbutyrate.
one or more of the polysaccharides such as
cellulose, pentosan, pectin, or starch as the
energy source, and most of the others
utilize hydrolytic products of these. How-
ever, a few species utilize only energy
sources such as glycerol (27) or, in the case
of methanogenic bacteria, C02-H2 or for-
mate (28). Relatively few ferment amino
acids and none utilize fatty acids such as
acetate or palmitate as an energy source.
A few selected features of 15 species repre-
senting 12 genera of representative rumen
bacteria are shown in Tables I-V. For
detailed references on these species, various
reviews are available (1, 2).
Nonmotile members of the genus
Bacteriodes are very numerous. The three
species shown (Table i) all produce suc-
cinate, acetate, and formate from carbo-
hydrate in a CO2-dependent fermentation
(21). Of interest is the fact that Bacteroides
ruminicola produces a small amount of
propionate via the acrylate pathway even
though succinate is the major fermentation
product. The B. ruminicola is the most
versatile member of this group in that it
ferments a large number of carbohydrates
and grows either with ammonia or pep-
tides, but not with amino acids, as the
main source of nitrogen (10). The other
two species require ammonia as the source
of nitrogen and utilize few energy
sources. Bacteroides succinogenes is prob-
ably the most actively cellulolytic bacterium
in the rumen (14) and is of further
interest in that it requires a straight-chain
acid such as n-valerate and a branched-
chain acid, isobutyrate or 2-methylbuty-
rate, for growth. Bacteroides amylophilus
ferments only starch and its hydrolytic
products such as maltose but not glucose
or other carbohydrates and has the greatest
biosynthetic capabilities of any known
rumen species. It has no requirement for
any carbon compounds or organic growth
factors other than the energy source and
CO2.Although the mineral requirements of
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Feature Succini-monas
Oval
+
+
None
S,A
(-CO2)
Forage-
grain
Helical
+
Few
S,A,F
(-CO2)
High
grain
Bulyrivibrio”
Curved
+
±
±
Many
B,A,F,L,EH2,CO2
9:
Many
Normal Flora-Rumen Bacteria 1445
few anaerobes have been studied, it is of
interest that both B. succinogenes (29) and
B. amylophilus (30) require Na+ for
growth. Although few nonmarine bacteria
are known to require Na+ and no other
anaerobes have been studied for this
feature, it is possible that many rumen
anaerobes require Na+ and that this
feature might serve as a convenient tool
for separation of these bacteria from non-
rumen anaerobes.
Some features of three of the most
important species of gram-negative rods
with monotrichous polar flagella are
shown in Table II. Two of these, Succi-
n ivibrio dextinosolvens and Succinimonas
amylolytica, are found among the domi-
nant species only when grain is included
in the diet and both produce succinate and
acetate as major products. Succinimonas
amylolytica ferments only starch and its
hydrolytic products. Succinivibrio dextrino-
solvens is often the most numerous bac-
terium when diets high in starch are fed.
Although it does not hydrolyze all com-
ponents of starch, all strains actively fer-
ment dextrin. The species Butyrivibrio
fibrisolvens is one of the most numerous
and ves-satile bacteria present in the rumen
and wide variations occur in features within
the group suggesting that more than one
species may be represented. This is the
major butyric acid-producing species in
the rumen and various strains ferment a
wide variety of energy sources including
compounds such as cellulose, starch, vari-
ous pentosans, pectin, and saponins. Re-
cent work with this species presented the
first demonstration of fermentation and
degradation of the heterocyclic ring struc-
ture of rutin and other bioflavonoids by
pure cultures of anaerobic bacteria (31).
It also hydrogenates trienoic and dienoic
18 carbon fatty acids to monoenoic acid.
In addition to butyrate, this organism may
either produce or fix acetate and produces
formate, lactate, ethanol, CO2 and H2 in
carbohydrate fermentation.
TABLE II
Some features of the more numerous rumen
anaerobic bacteria-gram-negative, motile
rods with monotrichous polar flagella
Shape
Energy sources
Glucose
Cellulose
Starch
Xylan
Others
Fermentation
products
NH3 from
amino acids
Animal diet
where num-
erous
Succini-vibrio
“Another unnamed organism (B-385-like) with
tufts of polar flagella and similar physiologically to
Butyrivibrio is often numerous, especially when the
rumen is somewhat acid. b Starch is not com-
pletely hydrolyzed but dextrin is fermented.S = succinate, A = acetate, F = forniate, B =
butyrate, L = lactate, E = ethanol, (-CO2) =
CO2 uptake.
Some features of other motile bacteria of
the rumen are shown in Table us. Seleno-
inonas ruminantium is a relatively large
crescentic bacterium with tufts of flagella
emanating from the concave side. Many
early workers thought it to be a flagellate
protozoan. It is very versatile in that it
ferments many carbohydrates and is one
of the major lactate- and amino acid-fer-
menting bacteria of the rumen. Its fermen-
tation of carbohydrate is unusual among
rumen bacteria in that propionic acid is
a major product, whereas most of the
rumen propionate is produced via succi-
nate production by many species and
succinate decarboxylation to propionate by
others (32). Lachnospira multiparus is
unusual in that it is a gram-positive curved
rod with lateral monotrichous flagella. It
is a very dominant member of the rumen
flora when legume pasture, high in pectin,
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Shape
Grain stain
Flagella
Spiro-
chete
Curved,
cres-
cent
Tuft
Concave
side
+
±
Many
P,A,L
CO2+
Many
Curved
+Mono-
trichous
Lateral
++
Few
A,F,L,E
CO2,H2
Legume
pasture
1446 Bryant
TABLE III
Some features of more numerous rumen
anaerobic bacteria-other motile bacteria
Feature Selenomonas Lachnospira Treponema
Energy sources
GlucosePectin
Lactate
OthersFerinen tation
products”
NH3 from amino
acids
Animal diet where
n imerous
= propionate, A = acetate, L =
F = formate, E = ethanol.
is fed and it appears to function primarily
in pectin fermentation. The rumen spiro-
chete, Treponema sp. (Borrelia sp.) is
almost always present but usually repre-
sents only about 4% or less of the total
cultured bacteria. It produces succinate as
a major product and appears to function
mainly in the fermentation of sugars pro-
duced during the hydrolysis of cellulose and
other polysaccharides by other species. This
was the first treponeme to be cultured in
a completely defined medium and was
shown to require large amounts of CO2
(16) and acids, such as valerate and iso-
butyrate, for growth (33).
Some of the major gram-positive non-
motile bacteria include Eubacterium rumi-
nan/in in, a bu tyrate-produci ng organism,
and an as yet unnamed strictly anaerobic
Lactobacillus sp. (Table iv). The latter
organism is homofermentative and pro-
duces n (-) lactic acid (34). It ferments
many sugars and is only numerous when
high grain rations or lush pasture is fed.
Under many conditions lactobacilli are
insignificant in the rumen.
The methanogenic bacteria are very
important members of the rumen fermen-
tation in that they utilize most of the
hydrogen and, possibly, formate, produced
by carbohydrate-fermenting bacteria, to re-
duce carbon dioxide to methane and to
- obtain energy for growth. They do not
None ferment other materials. Met hanoba cterium
ruminantium is the most numerous spe-
cies, although a new species, Met hano-
bacterium mobilis (28), also quite num-
+ erous under some condlitions, was recently? described. The M. ruminantium requires
ammonia as the main source of nitrogenMany
S,A,F,L and acetate and 2-methylbutyrate are essen-E tial as carbon sources (22).
- Table v shows some features of three of
the more important species of cocci. TheMany . -
ruminococci are usually present in large
numbers and are primarily concerned withlactate, fermentation of the polysaccharides, cellu-
lose, and xylan. It is of interest that most
strains do not ferment the monosaccharid!e
TABLE IV
Some features of the more numerous rumen
anaerobic bacteria-gram-positive
nonmotile rods
FeatureEubacierium
ruminan-hum
Laclobacillussp.
Meihano-baclerium
ru�ni-nanhium
Energy sources
Glucose + +Starch, cellulose, - - -
lactate
Xylan ± - -
H2-C02, formate - - +Fermentation B,F,L,A L CH4
products” CO2, H2
NH3 from amino - - -
acids
Animal diet where Forage Lush pas- Many
numerous ture, high
grain
“B = butyrate, F = formate, A = acetate,
L = lactate. The Laclobacillus sp. produces D (-)lactic acid.
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FeatureRumi-
nococcusalbus
0.7-1 .2
:1:
±
Few
A,F,E
CO2,H2
+
Many
Rumi-nococcus
flavefaciens
0.7-1.2
+
9:
±
Few
S,A,F
H2,
(-CO2)
+
Many
Pepto-siref’tococcus
elsdenii
1.2-2.4
+
+
+Few
A,P,B,V
C,C02,H2
+
High grain
Normal Flora-Rumen Bacteria 1447
hydrolytic products of these polysaccharides,
glucose, or xylose, although the disaccha-
rides, cellobiose, or xylobiose, are fer-
mented. This is presumably due to lack of
permeases for the monosaccharide (35).
The ruminococci require ammonia as the
main nitrogen source and most require one
or moie of the acids, isovalerate, isobuty-
rate, and 2-methylbutyrate for growth. The
two species are separated mainly on the
basis of fermentation products and chain
formation.
The major lactate fermenting species in
the rumen, other than Selenomonas rum i-
nantiu m, is Peptostreptococcus elsdenii
(Table v). This species is a major organism
in the rumen only when high grain diets,
or other diets that result in relatively low
pH, are fed. It is primarily functional in
lactate fermentation and is of particular
interest in that it is the major species
responsible for the production in the rumen
of larges- amounts of valeric and! caproic
acids from carbohydrate when high grain
diets are fed.
The question arises as to whether many of
the dominant species of rumen bacteria
are present in other habitats. Although
earlier workers often believed them to be,
for the most part, restricted to the rumen,
it is now evident that many identical or
closely related species are present in other
habitats. For example, ruminococci have
been isolated from the rabbit and guinea
pig cecum. Met han obacterium rum man-
tium is one of the dominant methane
bacteria involved in the anaerobic diges-
tion of sewage (20). It is the only methane
bacterium so far isolated from human
feces (36). Butyrivibrio sp. were isolated
from feces of humans, rabbits, and horses
(37). Both S. ruminantiuin and S. sputi-
genum from the human oral cavity have
recently been shown to be distinct species
(38), but it is possible that the guinea pig
cecal organism, Selenomonas palpitans,
which has not yet been isolated, and
selenonomads from the ceca of other
TABLE V
Some features of the more numerous rumen
anaerobic bacteria-gram-variable cocci
Size (�sm)
Chains
Energy sources
Glucose
Cellobiose, cellu-
lose, xylan
Lactate
OthersFermentation
products”
NH3 from amino
acids
NH3 and VFA re-
quired�
Animal diet where
numerous
“A = acetate, F = formate, E = ethanol, S =
succinate, P = propionate, B = butyrate, V =
valerate, C = caproate. b One or more of the
acids, isobutyrate, isovalerate, and 2-niethylbu-
tyrate ar� essential for growth.
rodlents are identical with S. ruminantium.
Studies on DNA homology may be neces-
sary in order to be certain that B. rumini-
cola is definitely different from some mem-
bers of the Bacteroides fragilis group
found in the intestinal tract of animals and
man. Further studies of habitats other than
the rumen and involving adequate isolation
techniques and definitive methods of iden-
tification such as those in use in the
Anaerobe Laboratory at Virginia Poly-
technic Institute (18) will materially aid in
solving this question.
Pathway of Carbohydrate Fermentation
Dr. Baldwin discusses the pathways of
fermentation in relation to energy genera-
tion (6); however, to close my discussion,
brief mention should be made of the path-
ways of carbohydrate fermentation in the
rumen in relationship to kinds of bactei-ia
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1448 Bryant
Polysaccharide - Fermenting Species
5: Sugar - Fermenting Species
C Lactate - Fermenting Species
D: Methanogenic Species
FIG. 1. Schematic presentation of kinds of bacte-ria, major end products (underlined), and majorextracellular intermediates (enclosed) formed in
fermentation of polysaccharides by rumen bacteria.
and extracellular intermediates. Figure 1
shows a simplified scheme of the fermenta-
tion of polysaccharides. Studies of fermen-
tations carried out by pure cultures and by
whole rumen contents indicate that poly-
saccharides are hydrolyzed to soluble oligo-
saccharides and sugars by polysaccharide-
hydrolyzing organisms. Then, these extra-
cellular intermediates are assimilated by
cells of these organisms and other sugar-
fermenting species and fermented with
production of some rumen end products
such as acetate, propionate, butyrate, and
CO2. However, these organisms usually also
produce end products that are further
metabolized by other species and, therefore,
are not rumen end products. These prod-
ucts, or extracellular intermediates, include
H2, formate, succinate, and lactate. The
formate is rapidly degraded to CO2 and
H2 by species of carbohydrate-fermenting
bacteria and methanogenic bacteria (39),
and the latter species very rapidly utilize
H2 to reduce CO2 to methane. Thus, one
almost never finds a significant concentra-
tion of formate or H2 in the rumen. The
succinate produced by many species is
rapidly decarboxylated with production of
propionate and CO2 by other species (32),
and lactate produced by some is fermented
by species such as P. elsdenii and S. rumi-
nantium with production of the acid end
products plus H2.
Pure culture studies suggested that lac-
tate and ethanol were important extra-
cellular intermediates in the rumen, but
studies of pooi sizes and turnover rates of
these compounds in whole rumen contents
indicate that very little of the carbohydrate
is fermented via lactate and essentially
none via ethanol under usual conditions
(1). Conversion via lactate increases if high
grain diets are fed, and under very unusual
conditions ethanol may be produced.
The discrepancy between the production
by pure cultures of much ethanol and
lactate and the fact that these compounds
are not significant intermediates in the
rumen under many conditions is probably
best explained by the fact that the methane
bacteria nearly always maintain a very low
partial pressure of H2. Most of the carbo-
hydrate-fermenting bacteria that produce
lactate or ethanol also produce acetate and
hydrogen. In pure culture fermentations,
hydrogen accumulates and the energetics
of electron flow from low-potential reduced
carriers, such as pyridine nucleotides and
ferredoxin generated in glycolysis and
pyruvate metabolism, to H2 become much
less favorable (40). Thus, in the pure
cultures more lactate and ethanol and less
acetate is probably produced in response to
an increased need for disposal of these
electrons. In the rumen and some other
anaerobic environments, the methane bac-
teria maintain a low partial pressure of
H2 that obviates the tendency for the
carbohydrate-fermenting bacteria to pro-
duce ethanol or lactate, and they probably
produce a larger amount of acetate. Acetate
formation from pyruvate generates adeno-
sine triphosphate, which provides energy
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Normal Flora-Rumen Bacteria 1449
1. HUNGATE, R. E. The Rumen and Its Microbes.
New York: Academic, 1966.
for growth of the organism, but lactate or
ethanol formation does not. Thus, the
probable importance of the methane bac-
teria in the fermentation becomes more
clear. They not only obtain energy for
growth by utilization of H2 but allow
hydrogen-producing bacteria to ferment
organic matter via pathways involving
more efficient energy generation (1, 40).
SUMMARY
The digestion of feeds, which usually
contain large amounts of cellulose and
similar polysaccharides, by the ruminant
animal is largely dependent on an intense
anaerobic microbial fermentation in the
rumen. Nonsporeforming anaerobic bac-
teria are mainly responsible for the fermen-
tation. The carbohydrates of the diet are
fermented mainly to acetate, propionate,
butyrate, CO2, and CH4, and these volatile
acids are a major source of energy for the
ruminant animal. Major extracellular in-
termediates, i.e., products of some micro-
bial species that are utilized by others in
the fermentation of polysaccharides include
oligosaccharides and sugars, H2, formate,
succinate and, under some conditions,
lactate. Microbial protein from the rumen
is usually the chief source of essential
amino acids for the ruminant regardless
of the type of protein in the diet and most
species of rumen bacteria can utilize
ammonia as the main nitrogen source. The
microorganisms also metabolize dietary
lipids and synthesize B-vitamins. The
methods of culture and some taxonomic,
nutritional, and metabolic characteristics
of some representative rumen anaerobic
bacteria are briefly discussed. These bac-
teria, in general, are better characterized
and their functions better understood than
those of any other anaerobic habitat.
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