Dietary fat and its influence on rumen microbiota. Part II

This process is considered as a detoxifying adaptation (Kemp et al. , 1984), and marginally contributes to the elimination of reducing equivalents produced by rumen fermentation (Lourenço, et al. 2010).

Biohydrogenation (BH) comprises several steps, depending on the USAFs, as well as several pathways, depending on diet and rumen environment (Griinari et al., 1998).

Protozoa encompass bacteria, and bacterial biohydrogenation can take place within protozoa (Jenkins et al., 2008) which explains their high concentrations of intermediate products (Devillard et al. , 2006).

In Vivo Studies

Beyond studies based on the selected isolates, in vivo trials have been carried out to evaluate the relationship between rumen bacteria and biohydrogenation. This has been done by  adding bacteria and quantifying their products, or by adding dietary supplements known to affect BH and measuring bacterial abundance.

As a rule

• In vivo studies of dietary fat inclusion have shown that there are no changes or reductions in total and/or major protozoan genera.

Other observations ….

• Hristov et al., 2012 observed that lauric acid (C12:0) strongly decreased protozoan counts compared to myristic (C14:0) and stearic (C18:0) acids.
• In a more general way, Oldick and Firkins (2000) showed that increasing the degree of unsaturation reduces protozoan count. However, they emphasized that it can be difficult to assess such change due to individual variation, which explains the inconsistency of their trials.

Observations on archaeal community

Methanobrevibacter ruminantium

Studies with pure strains of archaea adding organic acids or saturated fatty acids evidenced an inhibition of methane production by Methanobrevibacter ruminantium.

• Lillis et al. (2011) demonstrated that the addition of soybean oil in vivo altered the count of methanogenic archaeans but not their diversity. They hypothesized that these changes can be a consequence of an altered VFA profile(less acetate and butyrate producing H2, and more propionic), due to changes in the bacterial community.
• Hristov et al. (2012) proposed that changes in the archaeal community could be linked to a reduction in protozoa abundance when using high-fat diets.

About linoleic acid ….

• In pure cultures, linoleic acid (LA) can affect the growth of the fungus Neocallimastix frontalis (Maia et al. , 2007).
• Boots et al. (2012) confirmed the negative effect of LA on the order Neocallimastigales through in vivo trials. These researcher observed that the richness and diversity of these fungi are diminished with the addition of soybean oil.

• • • The inhibitory effect of oils on bacterial growth has been studied extensively in pure in vitro cultures of rumen strains (Maia et al., 2007). Focusing on bacteria that play a role in fibrolysis, amylolysis, and fat metabolism.

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Altogether these early studies on saturated and monounsaturated FAs emphasize that the effects of fats on rumen bacteria depend on: bacterial metabolism, FA unsaturation, and the geometric configuration of double bonds.

+ stronger for ALA than for LA

++strong for eicosapentaenoic long chain FA(EPA; cis-5, cis-8, cis-11, cis-14, cis-17-C20: 5 )and docosahexaenoic acids (DHA; cis-4, cis-7, cis-10, cis-13, cis-16, cis-19-C22: 6).

Similarly, ALA strongly increases the latency phase and decreases the growth rate of Propionibacterium acnesn (Maia et al., 2016).

Effects on bacteria in vivo

The effects of fat supplements were investigated in vivo, assigning bacteria to species level using quantitative PCR (Martin et al., 2016;  Vargas-Bello-Perez, et al., 2016) or, at genre level, using 16S rDNA pyrosequencing (Zened et al., 2013a;  Huws et al. , 2014).

In the latter, significant effects of fat were found in bacteria not yet cultured or not classified

These experiments were mainly based on the addition of oil, unlike most studies on pure strain cultures that used free FAs.

Overall, the effects were less than those observed with pure cultures. This could be due to the type of fat that was added or the fact that effects within the latency phase cannot be seen in vivo.

Changes in the rumen microbiota due to a higher concentrate proportion are much higher than the effects associated with addition of fats. It is also worth mentioning that some genera were affected differently by the addition of oil in low- and high-concentrate diets. This was especially true for Acetitomaculum, Lachnospira and Prevotella (Zened et al. 2011).

Among the genera of bacteria or species that were studied in several experiments, Fibrobacter and Ruminococcus were negatively affected in most cases, but the effects on Butyrivibio and Prevotella were highly variable.

The latter genera comprise many species with somewhat different functions, different metabolic pathways, and different sensitivities to FA in crops (Maia et al., 2007).

A decrease in the in vivo abundance  of a bacterial genus after a change in diet cannot be unequivocally interpreted as a direct effect of a diet, but could reflect a more global change in nutrient degradation and in the relationships between different microorganisms in the rumen.

How do FAs inhibit bacterial growth?

There are several hypotheses to explain the inhibitory mechanism of GGs in bacterial growth:

• Hypothesis nº 1 . Most lipids are associated with dietary particles in the rumen, and coating the dietary particles could impair adhesion, decreasing fiber degradation in the rumen (Devendra and Lewis 1974). However, this hypothesis is not consistent with the increase in the number of bacteria that attach to particles in the solid phase of the rumen when fat is added to the diet (Bauchart et al., 1986).
• Hypothesis nº2 .  Devendra and Lewis (1974) also proposed that dietary fat could decrease the availability of cations to bacteria, due to the formation of salts, which is consistent with the protective effects of calcium-rich substances on cellulose degradation when oil is added to the diet (Brooks et al.,  1954). However, it cannot explain all the negative effects of dietary fat.
• Hypothesis nº3 .  Devendra and Lewis (1974) also hypothesized that FA could exert direct toxicity on rumen bacteria, which is consistent with the incorporation of UFA into bacterial cultures (Bauchart et al. al. 1990). This toxicity could be due to an impediment in the passage of nutrients due to FAs attaching to the cell wall.

• A specific effect of cis double bonds could explain why most known bio initiator bacteria produce trans-C18:1, but do not reduce it further to stearic acid.
• Linoleic acid can alter cell integrity, but there is no relationship between this alteration and the level of inhibition of bacterial growth in different strains in the rumen, including fibrisolvens.

Conclusions and perspectives

The relationship between dietary lipids and rumen microbiota is dominated by the toxicity of UFAs in many microorganisms, especially fibrolytic bacteria.

• The genus Butyrivibrio is known to be strongly involved in the detoxification process.

Many recent studies suggest that the biochemical pathways are more complex and that the bacteria involved could be more diverse than previously believed several decades ago.

Practical applications involve both sides of this relationship.

• Dietary fat inclusion shapes the microbial community of the rumen, modulates the function of the rumen. As a result, a great amount of research in recent years has been devoted to reducing methane emissions, although the mode of action is not fully understood and could depend on the source of fat (Patra and Yu 2013).
• There are negative side effects on feed efficiency, such as reduced dry matter consumption (Beauchemin et al., 2009).

The most suitable options for shaping the rumen microbiota and its activity depend on many factors:

• production system
• economic conditions
• local regulations or specifications

However, to apply these different manipulations in the field, new data must be obtained in vivo under various dietary conditions with long-term studies because the resilience of the rumen microbiota or its adaptation to the degradation of plant compounds can alter the effects over time (Weimer 2015).

In addition, conducting new applied research on rumen fat metabolism makes it necessary to better understand which microorganisms, which enzymatic mechanisms and which interactions between microorganisms and between the microbiota and the host are involved.

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