The subtle (secondary) deficiency of methyl groups

One of the most important goals of bovine genetic selection is to improve milk protein production. Thanks to this selection and to the specific nutrition, today the high genetic merit (HGM) Holstein cows produce 3% casein during winter. This extremely high protein production is causing more and more fertility and immune problems to the animals. Bauman and Currie explained for the first time in their “Partitioning of nutrients during pregnancy and lactation: a review of mechanisms involving homeostasis and homeorhesis” (1980. J. Dairy Sci. 63:1514-1529) how nutrients are used by bovine metabolism and they made clearer some practical aspects of animal rearing.

Between calving and the new pregnancy, the cow’s udder has the metabolic priority over any other body function, except basal metabolism and body temperature maintenance. Genetic selection exasperated this udder “metabolic bullying” to improve milk, fat, and protein production, enhancing the physiological behavior of the animal to produce the highest milk quality (and quantity) to overcome its newborn needs. When the new pregnancy starts, the metabolic priority is for the uterus and the adipose tissue deposition to ensure the correct development and survival of the new fetus. The HGM cows are the ones with the highest milk production but also the highest weight loss before pregnancy and the highest weight increase after fertilization.

The udder takes the nutrients needed for milk production without hormone intermediations and this causes high productivity but also an unavoidable lack of nutrients in the fresh cows: the feed intake and rumen fermentation are not enough to supply the energy requirements. Negative energy balance (NEBAL) can be estimated through BCS variation and some milk and blood biomarkers. Among nutritionists and veterinarians, there is also the increasing awareness of a negative protein balance (NPB) that is more difficult to be calculated but causes huge damages to fertility and immunity. When cows experience strong metabolic stress, the amino acid dietary income is not enough and animals start to mobilize their own muscle proteins to overcome their needs. At the same time, they stop metabolic pathways that are not essential for life. The NPB can be evaluated through the echography of the muscle longissimus dorsii or through blood biomarkers such as 3-methylhistidine from the muscular protein (actin and myosin) catabolism, carnitine, and urea.

Fresh and non-pregnant dairy cows produce at least 45 kg of milk per day. Assuming 3.2% milk protein, the daily protein production is about 1440 g with 1100 g of casein. The cow needs 1.5 g of metabolizable protein (MP) to produce each gram of milk protein so that the udder requirement for such a daily production is about 2160 g of MP. It is estimated that between 2 weeks before calving and the first 5 weeks of lactation, the HGM bovines lose up to 115 kg of muscle tissue more or less corresponding to 21 kg of muscle protein.

Table 1: Different hormonal and metabolic “attitudes” in high and low genetic potential cows, during lactation or the dry period.
  Lactation start Dry period   Lactation start Dry period
  High production   Low production
Prolactin (ng/ml) 13.86 21.91 11.01 20.67
GH (ng/ml) 8.05 3.05 2.14 1.90
Insulin (µUI/ml) 24.45 22.60 24.45 22.60
Thyroxine (ng/ml) 30.70 38.40 39.80 55.40
Glucose (mg/ml) 0.639 0.753 0.628 0.735
NEFA (µEq/L) 358 161 217 213
BHBA (mg/ml) 0.125 0.103 0.080 0.102
Lactic acid (mg/ml) 0.076 0.081 0.091 0.101

Differences between the animal needs and the availability of the nutrients during the first 110-150 days of lactation lead to a secondary deficiency of nutrients. The primary deficiency is the lack of a nutrient if it is not included in the diet, while there is a secondary deficiency of that nutrient if it is available but in an insufficient quantity. Thus, HGM cows frequently experience secondary deficiencies of nutrients at the beginning of lactation. Highly digestible ingredients and rumen-protected feed additives (such as amino acids) help to reduce NEBAL and NPB.

Casein is the most important protein in milk and is about 77% of the total. There are different caseins, all composed of about 200 amino acids both essential and not. Italian data underlined that more than 25% of Italian Holstein cows produce < 2.9% milk protein during the firsts 75 days of lactation. At the same time, a lot of animals have a high genetic potential, but they produce low milk protein than expected. This is proof that, even if the udder and the milk synthesis is of priority importance, numbers of fresh cows have an amino acid deficiency.

The negative methyl donors balance (NMDB)

There are 20 amino acids among which about 50% are essential (EAA, they are not synthesized by the animal and they must be ingested with the diet) and the others are non-essential (NEAA, they are synthesized by the animal without risk of deficiencies). Lysine, methionine, valine, arginine, isoleucine, histidine, tryptophan, leucine, threonine, and phenylalanine are commonly defined as EAA, while glycine, serine, glutamic acid, cysteine, tyrosine, proline, alanine, aspartic acid, glutamine, and hydroxyproline are NEAA. Amino acids can be also divided among glucogenic, insulinogenic, and ketogenic ones.

Nowadays it is well documented that:

  • There is an excess of udder intake for EAA, in particular isoleucine, leucine, lysine, and valine;
  • Some of the dietary NEAA are not enough to overcome the requirement for milk protein production;
  • The udder synthesizes NEAA starting from the exceding EAA;
  • The use of lysine to produce NEAA can cause its deficiency;
  • Dietary extraction of proline, glutamic acid, and aspartic acid is lower than the udder requirements;
  • Proline and glutamic acid limit the protein synthesis even if they are synthesized by the udder;
  • The arginine taken by the udder is 2-4 times more than the one excreted in the milk;
  • Ornithine and citrulline are extracted by the udder, but not used in milk protein;
  • Milk contains 20-30% more phenylalanine and methionine than that extracted by the udder;
  • Phenylalanine, methionine, lysine, histidine, and threonine are the 5 amino acids essential for the mammary gland;
  • Arginine, valine, leucine, and isoleucine are extracted in higher quantities by the udder than their concentrations in milk protein;
  • The udder takes from the bloodstream enough proline, glutamate, and aspartate to overcome its needs.

Methionine is one of the best-known amino acids, is among the EAA, and is 5.5% of the total EAA in bovine milk and constitutes between 2.48% and 3.32% of casein. Cows take Met from the MP, which is composed of microbial protein and the dietary proteins that are undegraded in the rumen and reach the intestine, as well as from muscle protein. 50% of the total used Met comes from homocysteine remethylation. The enzyme methionine adenosyl-transferase synthesizes S-adenosyl-methionine (SAM) starting from methionine and ATP. SAM is a reactive molecule: methyltransferase removes the methyl group (CH3+) linked to the sulfur ion and donates it to another molecule (transmethylation). There are about 50 important metabolic reactions that use this transmethylation, involving molecules such as DNA, RNS, lipids, and proteins.

More than 30% of total methionine is used to produce choline. Met has also a pivotal role in apolipoprotein B and phosphatidylcholine synthesis. The Met requirements are strictly related to the dietary content of folates, vitamin B12, choline, and betaine. Figure 1 indicates the methyl groups metabolic pathways and highlights (the green circles) the interactions between particular molecules that can be used as rumen-protected feed additives for lactating dairy cows but also in HGM sheep, goats, and buffaloes.

Methyl groups have a relevant role in genetic expression. A number of genes are silenced through cytosine methylation, while histones acetylation makes part of DNA accessible to RNA transcription. These processes are influenced also by the environment. The whole process is called epigenetics and makes identical organisms phenotypically different one to the others without DNA modifications.

Figure 1: Methyl groups metabolic pathway. (Ruminantia, 2019).


Nutrition, metabolic alterations (NEBAL, NPB, NMDB), and metabolic pathologies are mainly related to the reproductive period. This temporal coincidence creates epigenetic modifications in the animals. The enzymatic defects that are inevitably accumulating in reared animals and the genetic modification induced by metabolic disorders have a sure negative impact on animal longevity and productivity. Antibiotics and hormonal therapies must be reduced to the indispensable, while clinical and function nutrition have (and will have) an increasingly important role.For more information: marketing@vetagro.comOriginal article here: part #1 and #2.