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A study of nutritional status of Finnish reindeer
(Rangifer Tarandus L.) in differents months

I. Composition and volume of the rumen microbiota

Liisa Syrjälä, Vappu Kossila and Helena Sipilä

Department 0/ Animal Husbandry, University of Helsinki

Received November 2, 1973

Abstract. The rumen microbiota were studied in free-ranging semi-domestic rein-
deer in Finnish Lapland under the nutritional conditions obtaining at two different
sampling times. Qualitative and quantitative investigations were made of the rumen
ciliate fauna and quantitative investigations of the rumen bacterial flora. The volume
coefficients for rumen ciliates obtained by Westerling (1970) and that for rumen
bacteria obtained by Warner (1962) were used to obtain an indication of the volume
of the rumen microbe mass in reindeer.

The rumen samples were collected in connection with the round-up and slaughter of
reindeer, being taken from 30 animals in December and 29 animals in March. The
reindeer slaughtered in December had normal access to food,but those slaughtered in
March had grazed on better pastures and received a supplementary feed of hay.

The total number of ciliate cells was over six times as high in March as in December,
the numbers being 1 182 900 and 188 300 per ml rumen contents, respectively. The
corresponding total numbers of bacterial cells were 9.65 x 109 in March and 6.65 x 109

in December. The reason for the statistically significantly (P < 0.01) higher numbers
in March than in December is probably the better nutritional conditions of the herd
slaughtered in March, not the time of the year.

The ciliate fauna consisted of 19 different species, although not all the species were
found in every sample. The percentage composition of the ciliate fauna did not vary
considerably between the two sampling times.

The volume of the total microbe mass constituted 8.2% of the rumen contents in
March and 1.9 % in December, the average being 5.1 %. The proportion of the ciliate
volume in the total microbe mass was clearly higher than that of the bacteria at both
sampling times: 7.2 times as high in March and 1.7 times in December, the average
being 4.7 times.

Since the importance of the rumen microbiota for the utilization of food
by ruminants was established, many investigations have been made of the
numbers and kinds of rumen microorganisms and the factors affecting them
(Warner 1965, Hungate 1966). The majority of the studies have been devoted
to the rumen microbiota of domestic animals, but some have also dealt with
semi-domestic or wild animals (Giesecke 1970). An extensive review of the

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535

rumen ciliate fauna of reindeer is given by Westerling (1970), but the rumen
bacteria of reindeer appear to have received less attention.

The purpose of this work was to study the quality and quantity of the
rumen ciliate fauna and the quantity of the bacterial flora of free-ranging semi-
domestic Finnish reindeer (Rangifer Tarandus L.) under the nutritional
conditions prevailing in two different months.

Experimental procedures

Sampling

The rumen samples studied were collected on 3.X11 1969 and 11.11 l 1970
from free-ranging reindeer in two areas of Finnish Lapland. The samples taken
in December were from animals in Lokka and those in May from animals in
Savukoski. The distance between these two areas is about 50 km. The samples
were obtained in connection with ordinary reindeer round-ups and slaughter.

The samples from the rumen were collected immediately after the animal
had been killed and the forestomachs removed. The wall of the rumen was
cut open and the contents were mixed thoroughly with a scoop, after which
25 ml of the rumen contents were measured with a glass measure and transferred
to a glass bottle containing 25 ml of 8 % formalin. As a rule, the rumen contents
were thickflowing, finely divided and fairly homogeneous, the colour being
gray-brown. No stratification was found.

Ciliate cell counts

A Wild-20 microscope with phase-contrast lighting was used for counting
the microbe cells.

The ciliate cells were counted and identified in a Fuchs-Rosenthal chamber,
as described by Westerling (1970). This chamber is divided into 16 rows of
16 square fields. The side of a square is 0.25 mm and the depth of the chamber
0.2 mm. The total capacity of the chamber is thus 3.2 mm2

. Usually the cells
in every second square row of the chamber were counted. Four counts were
performed on each rumen sample and thus the ciliates of 4 X (3.2 : 2) = 6.4
mm3 of diluted rumen contents were counted. The largest organisms were
counted in the whole chamber and half of the result was recorded. The original
rumen sample was kept in 8 % formalin solution. It was further diluted with
water to make the counting easier to perform. No staining was used, since the
ciliates were most easily identified in phase-contrast lighting without staining.
The total number of ciliates was calculated by adding the results for the sep-
arate species.

Bacteria cell counts

The bacterial cells were counted in a microscope whose ocular was marked
with a grid. The numbers of bacterial cells in a given amount of sample can



536

be calculated, if the area of the coverslip is known and also the area of the
grid squares at a certain magnification. The procedure was as follows (Syrjälä
1967): 0.01 ml of rumen sample, diluted with water and well mixed, was
pipetted on to the object slide and covered with a coverslip having an area of
24 X24 mm2 = 576 mm 2. The preparation was checked to see that the
cells were evenly distributed under the coverslip, and discarded if the distribu-
tion appeared unequal. The ocular grid consisted of 25 squares. When an
objective with a magnification of X 40 was used, the area of each square was
24.2 X 24.2 /i2

= 586 fx 2
. The area of the coverslip thus contained about

982 900 squares. The cells of 24 squares were counted on each preparation.
The choice of squares was intended to be as representative as possible of the
whole preparation. For instance, the first squares were always taken at the
same distance from the margin of the coverslip.

The amount of cells in 0.01 ml of sample was obtained by multiplying the
sum of the cells in 24 squares by 40 950 (982 900: 24). Three preparations
were made from each rumen sample for the bacteria cell counts. Only the
total number of cells was counted, no attempt being made to identify the
bacteria.

Determination of the volume of the microbe mass

The volume of the ciliate fauna was calculated by using the cell volume
coefficients for each cilliate species determined by Westerling (1970):

Dasytricha ruminantium 3.86 fi3 X 104

Entodinium simplex 1.07 *

E. dilobum 1.51 »

E. damae 0.33 *

E. anteronucleatum 3.60 *

E. quadricuspis 0.78 »

E. bicornutum 0.60 »

E. exiguum 0.26 »

E. longinucleatum 1.21 *

Diplodinium dogieli 17.57 o
D. rangiferi 151.73 »

Eudiplodinium impalae 6.89 »

E. spectabile 21.99 »

Ostracodinum magnum 70.87 »

O. obtusum 21.22 *

O. confluens 26,06 »

Enoploplastron triloricatum 8.49 »

Epidinium ecaudatum 12.09 »

E. gigas 51.92 »

In determining these coefficients, Westerling used the geometrical method
introduced by Schumacher (1962):

Length Width Thickness 3
Cell volume = x X X 712 2 2 4



537

For the bacterial cells the mean volume of 1 fis was used (Warner 1962).
The volume of the whole microbe mass in the rumen was taken as the sum of
the volumes of ciliates and bacteria.

Results and discussions

Number and kinds of ciliate and bacteria cells

Ciliate species. The ciliates found in the rumen of the reindeer
represented 19 different species (Table 1), which are the same as those found

Table 1, The mean number of ciliate (n x 103 ) and bacteria (n x 109) cells per ml rumen
contents and the percentage composition of the ciliate fauna.

Percentage composition
Mean numbers of fauna

December March December March

Number of samples 30 29
Ciliates, total 188.3 1 182.9 100 100

Dasytricha ruminantium 0.2 2,3 1.0 1.0 0.2 0.2
Entodinium simplex 10.1 104.4 5.4 8.8
E. dilobum 5.2 63.5 2.8 5.4
E. damae 8.1 92,6 4.3 7.8
E. anteronucleatum 88.0 428.3 46.7 36.2
E. quadricuspis 28.9 134.3 15.3 11.4
E. bicornutum 2.9 104.1 1.5 8.8
E. exiquum 6.1 50.8 3.2 4.3
E. longinucleatum 3.1 16.6 1.6 80.8 1.4 84.1
Diplodinium dogieli 8.0 21.3 4.3 1.8
D. rangiferi 0.2 9.8 0.1 4.4 0,8 2.6
Eudiplodinium impalae 6.7 38.3 3.6 3.2
E. spectabile 2.8 21.3 1.5 5.1 1.8 5.0
Ostracodinium magnum 1.4 3.1 0.7 0.3
O. obtusum 2.6 32,2 1.4 2.7
O. confluens 1.8 2.5 1.0 3.1 0.2 3.2
Enoploplastron triloricatum ... 2.1 18.1 1.1 1.1 1.51.5
Epidinium ecaudatum 3.9 12.9 2.1 1.1
E. gigas 6.2 26.5 3.3 5.4 2.2 3.3

Bacteria, total 6.65 9.65

by Westerling (1970). According to the review of Giesecke (1970) concerning
the rumen protozoa of different ruminant animals, species of the genera Dasyt-
richa and Enoploplastrum are not found at all in the rumen of reindeer. The
19 species found did not all occur in every animal. Especially in the samples
taken in December, some species were completely lacking. The more or less
casual absence of some ciliate species from the rumen of some animals in the
herd seems to be very common and may depend on chance or some temporary
fluctuation (Quinn et ai. 1962, Westerling 1970).



538

It has been shown on many experiments (Warner 1965) that even in
animals given the same dietary and environmental treatment the rumen
microbial populations may differ greatly both qualitatively and quantitatively.
When the same animal is kept under constant conditions, the rumen microbial
population may still show marked temporal variation. Besides changes in
the diet, starvation has an important influence on the rumen microorganisms.
During some days’ starvation the microorganisms die out at different rates
(Meiske et al. 1968, Warner 1965). This is important from the point of view
of the present study, since the samples were taken from animals rounded up
for slaughtering. During the round-up they are subject to extra exertion and
often have to go some days without any food, their only »food» being snow.

Total numbers of cells. Considerable variation was found
between the samples in the amounts of different micoroorganisms. The results
thus give information about the conditions in the herds rather than in the
individual reindeer.

The total numbers of ciliates and bacteria in the samples of March are
significantly (P<o.ol, tested by t-test) higher than in the samples of December
(Table 1). This is probably due more to the nutritional conditions than to the
time of year. Contrary to the usual situation the nutritional conditions were
more favourable for the herd slaughtered in March than for that slaughtered
in December. Before slaughtering the animals in the former herd had each
received a supplementary feed of about one kilogram of dried hay per day.
They were also kept throughout the winter on pasture that had been fenced
off and protected from grazing for the previous years. The reindeer herd
slaughtered in December had received no extra feed and its pasture had been
in continuous use for many years. If the two herds had been kept under nor-
mal nutritional conditions, the total numbers of ciliate and bacteria cells
could be expected to be higher earlier than later in the winter. It should be
mentioned here, that in winter 1969 a thick permanent snow cover was establish-
ed unusually early in Lapland, so that the reindeer had difficulty in obtaining
food as early as in November. Normally the food of reindeer becomes less
varied when winter arrives, which can be reflected in the rumen microorganisms.
When the animals have normal access to food, it has been noticed (Westerling
1970) that the total number of ciliates decreases from summer to winter.

Percentage composition of the fauna. The percentage
composition of the ciliate fauna did not show such large variations between the
two months as the amounts of cells (Table 1). Especially the representation of
different genera or subgenera in the total fauna seemed to vary less than that
of the single species. The small entodinia formed the highest proportion of the
fauna, on the average 82.5 %. This value is of about the same magnitude as that
obtained by Westerling (1970). He also found that Enlodinium anteronucle-
atum was the only one of the entodinia whose representation did not decrease
from summer to winter, which indicates that a lichen diet suits it better than
the other species of the genus. In the present study, the entodinia other than
E. anleronuclealum were better represented in the samples of March than in
those of December. E. anteronucleatum constituted 43 % of the numbers of
Enlodium in March and 58 % in December. This is understandable, because,



539

since the animals received hay in March, the proportion of the diet consisting
of lichens was presumably smaller then than in December.

The proportions of Diplodinium, Ostracodinium, Enoploplastron and Epi-
dinium in the ciliate fauna are of the same magnitude as in earlier experiments
(Westerling 1970), but the proportion of Eudiplodinium is less than half
that of the values obtained by Westerling.

Volume of the microbe mass

In the volumes of both ciliates and bacteria there were found statistically
significant (P<0.01) differences between the two months. The total volume
of ciliates per ml rumen contents (Table 2) was about 6 times as high in March

Table 2. The mean volume of ciliates {u3 x 10*) per ml rumen contents and its percentage
distribution by genera or subgenera.

Volume Percentage distribution
December March December March

Ciliates, total 12 354 72 365 100.0 100.0
Dasytricha 8 89 0.1 0.1
Entodinium 3 678 19 805 29.8 27.4
Diplodinium 1 709 18 612 13.8 25.7
Eudiplodinum 1 077 7 323 8.7 10.1
Ostracodinium 2 013 9 681 16.3 13.5
Enoploplastron 178 1 537 1.4 2.1
Epidinium 3 691 15 318 29.9 21.2

as in December. The largest proportions of the ciliate volume in both months
related to the genera Entodinia and Epidinium. In March the genus Diplodi-
nium was also well represented, consitituting 25.7 % of the volume, whereas
in December it amounted to only 13.8 %.

The proportion of the total volume of the microbe mass in the rumen
contents (Table 3) was found to be 8.2 % for samples in March and 1.9 % in

Table 3. Microbe mass volume as % of rumen contents

December March Average

Ciliates 1.2 7.2 4.2
Bacteria 0.7 1.0 0.9
Total 1.9 8.2 5.1

December, the average being 5.1 %. These values are of about the same mag-
nitude as those obtained for other ruminants (Warner 1965), but are lower



540

than the values for rumen ciliates alone reported for reindeer by Westerling

(1970). He found the total volume of rumen ciliates in reindeer to be about
14 % of the rumen contents which is almost 100 % more than that demonstrated
in other hosts and did not observe any variations attributable to diet or season
when the animals had normal access to food. This indicates that the magnitude
of the ciliate mass in the reindeer forestomachs does not diminish when the
animal changes from a protein-rich summer diet to a winter diet poor in protein.

The microbe mass in the rumen is a good index of rumen microbial activity
and the protein economy. According to several estimates more than half of
the rumen nitrogen is in the form of microbial cells (Gray et al. 1953, Weller
et al. 1958). The rumen microbe mass can be very important as a protein
reserve especially in wild ruminant animals such as reindeer, whose nitrogen
intake can be very unstable in the winter. This explains, why the volume
occupied by microorganisms in the rumen of wild animals has been observed
to be notably high.

The proportion of ciliates in the total volume of the microbe mass is clearly
higher in both months than that of bacteria: 7.2 times as high in March and
1,7 times in December, the averege being 4.6 times. In domestic ruminants
the ciliate mass has been found to be be roughly equal to that of the bacteria
(Abou Akkada 1965). The present results indicate that the amount of ciliates
decreases more than that of bacteria when the food supply diminishes or the
food becomes less varied. This relatively faster decrease in the representation
of ciliates can be explained by the fact that they are more sensitive than the
bacteria to any changes in the environmental and nutritional conditions
(Hungate 1966). However, apart from some exceptions, the percentage
composition of the ciliate fauna (Tables I—2) was about the same in the two
months. The changes in question thus relate to the total numbers or volume
of the ciliate fauna.

REFERENCES

Abou Akkada, A. R. 1965. The metabolism of ciliate protozoa in relation to rumen function.
Physiology of digestion in the ruminant, 335 345. Ed. Dougherty. R. W., Washington.

Giesecke, D. 1970. Comparative microbiology of the alimentary tract. Physiology of diges-
tion and metabolism in the ruminant, 306 318. Ed. Phillipson, A. T., Newcastle
upon Tyne.

Gray, F. V., Pilgrim, A. F. & Weller, R. A. 1953. Conversion of plant nitrogen to microbial
nitrogen in the rumen of the sheep. Nature 172: 347 348.

Hungate, R. E. 1966. The rumen and its microbes. 533 p. New York.
Meiske, J. C., Salisbury, R. L,, Hoeter, J. A. & Luecke, R. W. 1958. The effect of starva-

tion and subsequent refeeding on some activities of rumen microorganisms in vitro.
J. Anim. Sci. 17: 774-781.

Quinn, L. Y., Burroughs, W. & Christiansen, W. C. 1962. Continuous culture of ruminal
microorganisms in chemically defined medium. 11. Appi. Microbiol. 10: 583 592.

Schumacher, E. 1962. fiber die Wirkung einiger Sulfonamide und Antibiotika auf die Infusorien
und die Gärgasbildungen im Panseninhalt des Rindes. Schweiz. Arch. Tierheilk. 104:
491-518.

Syrjälä, L. 1967. Märehtijäin ruoansulatuksen mikrobeista ja näiden kettoaineita hajoittavasta
toiminnasta. M.Sc. Thesis. Dept. Microb., Univ. Helsinki. 100 p.



541

Warner, A. C. I. 1962. Some factors influencing the rumen microbial population. J. Gen.
Microbial. 28:129-146.

» 1965. Factors influencing numbers and kinds of micro-organisms in the rumen. Phys-
iology of digestion in the ruminant, 346 359. Ed. Dougherty, R. W., Washington.

Weller, R. A., Gray, F. V. and Pilgrim, A. F'. 1958, The conversion of plant nitrogen to
microbial nitrogen in the rumen of the sheep. Br. J. Nutr. 12:421—429.

Westerling, B. 1970. Rumen ciliate fauna of semi-domestic reindeer (Rangifer Tarandus L.)
in Finland: Composition, volume and some seasonal variations, Acta Zoolog. Fenn.
127; 1-76.

Selostus

Porojen ravitsemustilaa koskeva tutkimus
I. Pötsimikrobiston koostumus ja volyymi

Liisa Syrjälä, Vappu Kossila ja Helena Sipilä

Helsingin yliopiston kolieläintieteen laitos

Tutkimuksessa selvitetään luonnonvaraisena elävien porojen pötsin alkueläinten määrää
ja laatua, bakteerien määrää sekä mikrobimassan osuutta pötsin tilavuudesta kahtena eri
ajankohtana. Näytteet pötsistä otettiin erotuksen yhteydessä teurastetuilta poroilta, joulu-
kuussa 1969 30 eläimeltä ja maaliskuussa 1970 29 eläimeltä. Joulukuussa teurastetuilla poroilla
oli ollut normaalit luonnonvaraisten porojen ruokinnalliset olosuhteet, kun taas maaliskuussa
teurastetut porot olivat olleet paremmilla laitumilla sekä saaneet muutamia päiviä ennen teu-
rastusta lisärehuna heinää.

Alkueläinten kokonaislukumäärä oli maaliskuussa yli 6 kertaa suurempi kuin joulukuussa,
vastaavien lukuarvojen ollessa 1 182 900 ja 188 300 per ml pötsin sisältöä. Alkueläimistön
prosenttinen jakautuminen eri lajeihin oli sitävastoin lähes samanlainen kumpanakin kautena.
Alkueläinlajeja tavattiin yhteensä 19. Bakteerien kokonaislukumäärä oli maaliskuussa 9.65 x
10® ja joulukuussa 6.65 X 10®. Syy merkitsevästi (P < 0.01) korkeampiin mikrobisolujen
lukumääriin maaliskuussa verrattuna joulukuuhun johtuu todennäköisesti laumojen erilaisista
ruokinnallisista olosuhteista eikä niinkään vuodenajasta. Koko mikrobimassan osuus pötsin
sisällön tilavuudesta oli maaliskuun näytteissä 8.2 % ja joulukuun näytteissä 1.9 % eli keski-
määrin 5.1 %. Alkueläinten osuus mikrobimassan tilavuudesta oli selvästi suurempi kuin
bakteerien osuus; maaliskuussa 7.2 kertaa ja joulukuussa 1.7 kertaa suurempi, keskiarvon ol-
lessa 4.7.