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Defaunation of the rumen

J.P. JOUANY

Introduction

Protozoa represent approximately 50 % of the microbial biomass in the rumen. Their presence has been shown to be important but not essential for the ruminant. Because interactions between microbes in the rumen are common, the role of protozoa is difficult to study separately, in a pure culture for instance. Moreover, because of the complexity of the rumen ecosystem, scientists are obliged to simplify the system to understand the specific role of certain microbes or the microbial interactions in the anaerobic digestive processes. These conditions were reached in gnotoxenic animals which harbour controlled species of bacteria and fungi. Similar models were made in France in the 1970’s with sheep inoculated with only one genus of protozoa (Polyplastron or Entodinium or Isotricha spp.) in association with a mixed population of bacteria and fungi.

Elimination of all the protozoa (defaunation) from the rumen is a standard method of studying their overall effect. Defaunation is also an essential step when growing in vivo a clone from a single cell of protozoa, or a mixed but controlled population in the rumen. Such a model of rumen inoculated with only one genus of protozoa has been used in our laboratory to study the specific role of the different genera and is a novel approach to understand the real contribution of protozoa to rumen digestion.

1. Critical study of the different methods of defaunation

In early studies, Becker et al. (1929/1930) showed that it was possible to eliminate protozoa from the rumen. Since then, many trials have been carried out to develop a simple and efficient method of defaunation. As previously established (Jouany and Ushida, 1990), such a method must

  • - eliminate protozoa completely
  • - be harmless to the animals
  • - have no drastic effect on bacteria or fungi

J.P. Jouany ed. : Rumen microbial metabolism and ruminant digestion.
INRA Editions, Paris, 1991, 239-261

Achieving these conditions together is far from easy because protozoa are particularly well adapted to the rumen habitat.

Different treatments have been suggested for defaunation of the rumen

  • - Isolation of animals a few hours after birth (Abou Akkada and El Shazly, 1964; Borhami et al., 1967; Chalmers et al., 1968; Eadie et Gill, 1971; Williams and Dinusson, 1973).
  • - Chemical defaunation associated or not with a 24- to 72-hour starvation period. Different chemicals have been used: copper sulphate (Becker et al., 1929/1930; Christiansen et al., 1965), dioctyl sodium sulphosuccinate (Abou Akkada et al., 1968; Orpin, 1977; Demeyer et al., 1982), alcohol ethoxylate or alkanates (Bird and Leng, 1978, 1984; Bird et al., 1979), calcium peroxide (Demeyer, 1982), coconut oil (Sutton et al., 1983; Newbold and Chamberlain, 1988), linseed oil (Ikwegbu and Sutton, 1982; Broudiscou and Lassalas, 1990) and soya oil hydrolysate (Broudiscou et al., 1988).
  • - Emptying the rumen, washing carefully the rumen mucosa, and treating the rumen contents before they are reintroduced. Heating the rumen contents to 50°C for 15 minutes kills the holotrich ciliates (Eadie and Oxford, 1957); freezing eliminates all the protozoa (Jouany and Sénaud, 1979a).


It is not judicious to believe that the only difference between controlled and treated animals is the presence or the absence of protozoa in the rumen. The method used to defaunate must be taken into account when analysing the disagreements between authors on the effect of protozoa. The behaviour of the other components of the ruminal ecosystem (bacteria and fungi) during treatment, should also be considered.


For the “isolation method”, it is advised that young animals stay with their dams for 72 hours before being isolated, to allow the inoculation of the rumen with strictly anaerobic bacteria (cellulolytic and methanogenic bacteria) (Fonty et al., 1987, 1988). Anaerobic fungi usually appear later, during the second week of life, approximately at the same time as protozoa. Thus, it is likely that animals with a defaunated rumen reared in strictly isolated conditions according to this method harbour no fungi.


Chemical defaunation should be carried out with great care and only for short periods because the threshold of toxicity for protozoa is close to that for animals. It has been shown that chemical agents have a direct toxic effect on bacteria and probably on fungi too (Eadie and Shand, 1981), which suggests that the observed effect of defaunation involves more than the elimination of protozoa.


To cancel out the deleterious effects of defaunation, it is strongly advised to reinoculate the rumen with thawed rumen contents, because freezing kills protozoa and preserves fungi and bacteria (Hillaire and Jouany, 1990).


A scientific comparison of different defaunating methods is difficult because efficiency is largely influenced by the experimenter’s familiarity with the technique. Our experience has shown that some faunated animals are difficult to defaunate. This might be due to a contamination from the omasum (protozoa in this organ not being submitted to the treatment). We also observed that defaunation is systematically more difficult for some experimenters than others, even if they apply the same technique to the same animals in the same laboratory!

2. Critical study of the experimental schedules used to test the effect of defaunation

Comparisons of faunated and defaunated animals are generally made on a small number of animals. Often, the same animals are used as controlled and treated...