To My Quyen, Nguyen Khanh Thuan, Nguyen Phuc Khanh, Nguyen Thanh Lam*

1. Introduction

Hypocalcemia is a metabolic disease, commonly seen in crossbred cows due to milk fever. It is characterized by inability to stand, general muscular weakness, circulatory collapse, depression and loss of consciousness (Hamali, 2008). Apart from this, it has been also reported in cows during ephemeral fever, coliform mastitis, and after parenteral administration of aminoglycosides. Hypocalcemia reduces rumen and abomasal motility increasing the risk of abomasal displacement, reduces feed intake, increases the risk of mastitis, reproductive performance, impairs immune function (Goff, 2008a; Kimura et al., 2006). 

2. Calcium homeostatic mechanisms

Ca is essential for life in animals. It is involved in many fundamental biological processes in the body, such as bone formation, muscle contraction, nerve transmission, and blood clotting, and it serves as a second messenger regulating the actions of many hormones. Therefore, it is of major importance that Ca concentration is regulated within a narrow range. In mammals, this process involves a coordinated effort among the hormones 1,25-dihydroxyvitamin D3, parathyroid hormone (PTH), and calcitonin. Homeostatic control of Ca concentrations in blood is so strong that variations are small and do not reflect dietary intake at all (Whitaker, 2000).

The skeleton of a 600-kg cow contains approximately 8.5 kg of Ca. There are 3 g Ca in the plasma pool and only 8 to 9 g Ca in all the extracellular fluids of a 600-kg cow. Blood Ca in the adult cow is maintained between 2.1 and 2.5 mmol/L (8.5 and 10 mg/dL) and is slightly higher in young animals. About 50% of the blood Ca is bound to proteins such as albumin, less than 10% is in mineral complexes with inorganic phosphates and the remainder exists in the ionized form. The ionized Ca concentration is the biologically active form of the Ca in blood and is most important for immediate metabolic function. During acidosis, larger numbers of protons compete with Ca (and with other cations) for binding to anionic sites of plasma proteins such as albumin. This drives more protein-bound Ca into solution, thereby increasing the ionized Ca concentrations. Conversely, alkalosis decreases the ionized Ca concentrations (Thrall et al., 2012). Cows that are alkalotic from upper gastrointestinal obstruction or other conditions may have normal total Ca while exhibiting clinical signs of hypocalcemia. Under acidic conditions, the ionized portion of Ca in the blood is closer to 48%; under alkaline conditions, it is closer to 42% ionized. The final 3% to 7% of Ca in blood is bound to soluble anions, such as citrate, phosphate, bicarbonate, and sulfate. If total serum proteins are greatly reduced (hypoalbuminemia), it is possible to have low total Ca in the blood and relatively normal levels of ionized Ca in the blood. Appropriately measured ionized Ca is the gold standard to evaluate physiologically active forms of Ca in a patient. Adjusted formulas were suggested to correct Ca concentrations in regard to albumin or total protein variations in dairy cows, when total Ca is measured (Seifi et al., 2005). However, serum total Ca concentration has routinely been the primary source for evaluation of Ca abnormalities in dairy cows. 
Dairy cows in early lactation producing colostrum (containing 1.7-2.3 g Ca per kg) or milk (containing 1.2 g Ca per kg) typically secrete 30 to 40 g of Ca each day. The total amount of Ca in plasma and extracellular fluids is estimated to be 12 g. Thus, body experiences negative Ca balance with initiation of lactation in dairy cows. Blood Ca concentration remarkably declines in dairy cows around calving, with the lowest concentrations occurring about 12 to 24 hours after calving. Blood samples obtained at this time can reveal the extent of hypocalcemia experienced by a dairy herd (Goff, 2014).

The drain of Ca during early lactation (30–50 g/day) represents a significant increase in Ca demand needed for late fetal growth (10-15 g/day). During the dry period, the supply of Ca through the diet is usually more than requirements of dam and fetus. Thus, passive Ca transfer from intestine is adequate to maintain homoeostasis without activating the Ca mobilization system, which is usually not activated until parturition. The main site for Ca absorption is assumed to be the small intestines, at least at moderate Ca intakes. The capacity for cows to absorb Ca through the rumen and abomasum is uncertain (Holler et al., 1988).

The demand of the mammary gland for Ca often exceeds the ability of the cow to replenish the plasma Ca pools, resulting in an acute decrease in the plasma Ca concentration in most cows. To maintain Ca homeostasis after calving, at the start of lactation, Ca-maintaining homeostatic mechanisms are activated. These mechanisms include: increased Ca reabsorption in the kidneys, increased Ca absorption in the intestine and Ca withdraw from bone (Goff, 2008a). Two hormones, 1,25 dihydroxy cholecalciferol (1,25(OH)2 D3) and parathyroid hormone (PTH) are involved in each of these processes (Figure 1).

3. Aetiology 

Hypocalcemia has a number of causes, including

• Age 
• Metabolic alkalosis 
• Hypomagnasemia
• High dietary Ca
• High dietary phosphorus 

3.1 Age 

Age has a profound effect on susceptibility of dairy cows to hypocalcemia. Older cows are affected by hypocalcemia more common and more severe than young cows. The Ca output, however, does not explain the increase in prevalence of hypocalcemia within multiparous cows with increasing age as their colostrum yield is not different. The age and parity-associated susceptibility might be related Ca homeostatic mechanisms. With increasing age, the hemostasis process is impeded and results in moderate to severe hypocalcemia. It has been assumed that the number of vitamin D receptors in intestines decline with increasing age. In addition, as animals age increase, the number of receptors for PTH on target tissues decline (Hanai et al., 1990)

3.2 Metabolic alkalosis

Current evidence suggests that milk fever and hypocalcemia may occur in cows as a result of excessive dietary cations. The typical ration of dairy cows are rich in cations (primarily potassium and sodium) and has lower amounts of anions (chloride and sulfur). These cations induce a metabolic alkalosis in the cow that impairs Ca homeostatic mechanisms via attenuated responsiveness of tissues to PTH. It has been shown that cows in a more acidotic state, which can be measured by the pH in the urine, have a decreased risk of developing milk fever (Seifi et al., 2004). 

In cows fed diets rich in cations, a greater number of positively charged cations enter the blood than negatively charged anions and results in an electrical disparity. To restore electroneutrality to this positively charged blood, a positive charge in the form of a hydrogen ion (H+) is lost from the blood compartment and the pH of the blood is increased. Adding readily absorbable anions to the diet increases the total negative charges in the blood, allowing more H+ to exist and a decreased blood pH, restoring tissue sensitivity to PTH (Goff, 2004). 

Under normal conditions, PTH released in response to hypocalcemia interacts with its receptor, located on the surface of bone and kidney cells. This stimulates G-proteins and adenylate cyclase resulting in production of cyclic AMP, which acts as a second messenger within the cytosol of target cells. This initiates mechanisms such as bone Ca resorption and renal production of 1,25(OH)₂ D3 to restore blood Ca concentration to normal levels. In alkalotic conditions induced by high cationic diets, the shape of the PTH receptor protein is changed so that it is less able to recognize and bind PTH, resulting in failure to activate the cell by producing cyclic AMP. Furthermore, Mg is required for full function of the adenylate cyclase complex (Goff, 2008a). 

3.3 Hypomagnasemia 

Hypomagnesemia affects Ca metabolism by reducing PTH secretion in response to hypocalcemia, and by reducing ability of PTH stimulated cells to produce cyclic AMP, resulting in failure to activate the target tissues to PTH. On the contrary, serum Ca and magnesium concentrations are negatively associated. Cows suffering from hypocalcemia have higher serum magnesium concentration. In a period of low serum Ca concentration, PTH is secreted into the blood. PTH secretion raises the threshold for renal magnesium excretion, resulting in a higher serum magnesium concentration (Martín-Tereso and Martens, 2014). 

3.4 High dietary Ca 

Before calving, the approximate daily requirement for Ca is only 30 g, comprising 15 g in fecal and urinary loss and 15 g to fetal growth. When supplying Ca far in excess of the daily requirements at dry period, the passive transfer of Ca is sufficient to overcome the needs of the cow and the fetus. Therefore, active hemostatic mechanisms of absorption and resorption of Ca become depressed. As a consequence, at calving when sudden massive demands for Ca occur, the cow is unable to rapidly return to hemostatic mechanisms and is susceptible to severe hypocalcemia until these mechanisms can be activated, which may take several days (Constable et al., 2016). 

3.5 High dietary phosphorus 

In dry cows, high dietary levels of phosphorus (more than 0.5% dry matter intake) increase the serum level of inorganic phosphorus, which has inhibitory effect on the renal enzyme (1a-hydroxylase) that catalyzes the conversion of vitamin D into its active form (1,25(OH)2 D3) and thereby predisposes cows to hypocalcemia. High dietary phosphorus has also been reported to have a negative affect on intestinal magnesium absorption, which further makes periparturient cows susceptible to hypocalcemia (Schnewille et al., 1994).

4. Epidemiology 

The overall incidence of milk fever found from a national dairy study in United States was 5%. Other field studies reporting incidence of milk fever from 1977 to 2007 found that the incidence in 10 North American studies was 3.45% (range 0–7%), in 10 European studies it was 6.17% (range 0–10%), and for 10 Australasian studies it was 3.5% (range 0–7%). Clinical milk fever was prevalent in 1, 4, 6, and 10% of first, second, third, and ≥ fourth lactation cows, respectively. 

Prevalence of clinical milk fever in a recent German study was higher (13.4, 15.0, and 21.7% for fourth, fifth, and sixth parity, respectively). It is assumed that preventive strategies are more common in the United States. German farmers favored oral Ca supplementation more than anionic salts as a preventive strategy. They supplemented Ca subcutaneously or orally in 46.1% of the herds and only 8.7% of the herds used anionic salts to prevent hypocalcemia. Based on the US agriculture ministry report, 68.9 and 27.6% of the herds used Ca products and anionic salts, respectively. The same report indicated that 20.7% of heifers and 27.6% of cows were fed anionic salts (USDA, 2016). The incidence of subclinical hypocalcemia – blood Ca values between 2 and 1.38 mmol/L (8 and 5.5 mg/dL) during the periparturient period – is around 50% in older cow. In a recent German study, 47.6% of multiparous cows suffered from subclinical hypocalcemia (less than 2 mmol/L) within 48 h after parturition. This finding is in agreement with previous studies. Subclinical hypocalcemia increased with age and was present in 41%, 49%, 51%, 54%, and 42% of 2nd–6th lactation cows, respectively (Figure 2) (Reinhardt et al., 2011).

5. Pathophysiology 

During the dry period, the demand of calcium is relatively low. Hence, intestinal absorption and bone resorption of calcium is relatively inactive during this time. Parturition is accompanied by sudden increase in calcium sequestration for the production of milk. The calcium requirements rise to 2 to 5 times that of dry period. This daily calcium out flow through milk will not be matched with decreased plasma calcium. This places major strain on calcium homeostatic mechanism. Decreased plasma calcium cause hyperexcitability of nervous system and weakened muscle contractions, which result in both tetany and paresis. The fall in blood calcium level stimulates the calcium homeostatic mechanism to improve intestinal absorption and bone resorption. Therefore, the decrease in plasma calcium causes a compensatory increase in parathyroid hormone and calcitriol, but it takes time for both to exert their full effect (Harris, 1981). Bone calcium mobilization by parathyroid hormone takes at least a week and improved efficiency of calcium absorption by influence of calcitriol takes a day or two. So, nearly all animals at parturition develop hypocalcemia, but high yielders tend to develop milk fever. The pathogenesis of the disease is more associated with the action of PTH that is responsible for regulation of calcium homeostatic mechanism. Hypocalcemia affects muscular contraction mainly in three ways. Firstly, calcium has a membrane stabilizing effect on the peripheral nerves. So hyperesthesia and mild tetany is seen in early stages of milk fever. Secondly, calcium is required for the release of acetylcholine at the neuromuscular junction. The inability to release acetylcholine causes paralysis by blocking the transmission of nerve impulse to the muscle fibers. Thirdly, calcium is directly required by muscle cells for contraction (Iggo, 1970). There is decreased contractility of cardiac muscle and lowered stroke volume cause reduction in arterial blood. Then this reduced peripheral perfusion resulted in hypothermia and depression of consciousness. Hypocalcemia also reduces gastrointestinal (GIT) function. Serum calcium content below 5 mg/dL reduces abomasal motility and rumen function and thus reduces energy balances, which is manifested as elevated blood nonesterified fatty acid (NEFA). Periparturient cows also experience immune suppression. Intracellular calcium signaling is a key early feature in immune cell activation, so the increased demand for calcium in these cows affect intracellular calcium stores of immune cells and leads to the immune suppression (Kimura, 2006).

Ca is necessary for proper function of a wide variety of systems in the body from structural functions such as bone and other tissues to intracellular processes as a second messenger. Extracellular Ca is necessary for muscle contraction, nerve impulses, blood clotting, and is a component of milk and bone. Intracellular Ca is involved in second messenger systems for a wide variety of processes. Hypocalcemia is considered as a gateway disease (Figure 3) and predisposes the cow to various metabolic and infectious disorders in early lactation such as metritis, mastitis, abomasal displacement, and reproduction disturbances (Goff, 2008b). 

Immune suppression 

It is well documented that nearly all dairy cows experience some degree of immune suppression during the transition period. Contributing factors for immune suppression in transitional period consist of decreased polymorphonuclear leukocytes, glycogen stores, decreased blood Ca concentration and increased non-estrified fatty acids (NEFA) and ß-hydroxyl butyrate (BHBA). Ca is critical for proper immune cell function, which is very important in transition dairy cows. In a study by Martinez et al. (2012), numbers of neutrophils were reduced and their ability to undergo phagocytosis and oxidative burst was impaired in cows affected by hypocalcemia, which might in part explain the increased risk for infectious diseases . Hypocalcemia is associated with decreased intracellular Ca stores in peripheral mononuclear cells. This is the cause of a blunted intracellular Ca release response to an immune cell activation signal. Kimura et al. (2006) concluded that intracellular Ca stores decreases in peripheral blood mononuclear cells before parturition and development of hypocalcemia. This decrease contributes to periparturient immune suppression (Kimura et al., 2006). Ca also regulates cell polarity, which is required for directional cell killing, and it is also involved in the migration of leukocytes toward chemokines in the area of inflammation (Gallo et al., 2006). 

Feed intake and weight loss 

It has been shown that cows with subclinical hypocalcemia have impaired rumen and abomasum motility and depressed feed intake. This reduction in ruminal and abomasal motility will likely cause a reduction in feed intake and increased weight loss in early lactation. Therefore, hypocalcemia may well exacerbate negative energy balance in cows that are already underfed (Mulligan and Doherty, 2008). 

Ketosis 

Hypocalcemia has been attributed to the occurrence of ketosis. Rodríguez et al. (2017) showed that hypocalcemic cows demonstrate 5.5 greater odds of having ketosis than normocalcemic cows (Rodríguez et al., 2017). The exact mechanism is unknown, however, the hypocalcemia impact on feed intake and resulting negative energy balance may be a factor in promoting of ketosis. Cows with naturally occurring hypocalcemia at parturition and experimentally induced hypocalcemia had elevated concentrations of NEFA and BHBA as indicators of increased lipid mobilization (Martinez et al., 2014). 

Lipolysis 

Ca is also important in adipocytes for regulating lipid metabolism and triglyceride storage. In studies with rat and human adipocytes, increased intracellular Ca has been shown to have antilipolytic effects. It can be speculated that hypocalcemia may deplete adipocyte Ca stores, resulting in increased lipolysis (Seifi and Kia, 2017). 

Abomasal displacement 

It has been shown that hypocalcemia increases the risk for displacement of abomasum. Abomasal atony due to hypocalcemia seems to be a logical risk factor for abomasal displacement. Hypocalcemia may reduce abomasal tone and result in gas accumulation. It was reported that 82% of the cows with displaced abomasum had Ca values equal or less than 2.0 mmol/L in the first week after calving. There is also a report from one herd that subclinical hypocalcemia at calving was a risk factor for left displacement of abomasum. Seifi et al. (2011) showed that the odds of the development of displacement of abomasum were 5.1 times greater in cows with serum Ca concentrations equal or less than 2.3 mmol/L in the first week post-partum. Reduced Ca concentrations have been associated with a reduction in rumen and abomasal motility, which in turn is thought to increase the risk of abomasal displacement. It is likely that Ca concentration is an indicator of inadequate dry matter intake, which most likely contributes to the development of displacement of abomasum (Chapinal et al., 2011). However, Leblanc et al. (2005) did not find a direct relationship between Ca concentrations and left displacement of abomasum incidence and suggested that subclinical hypocalcemia may be a function of decreased feed intake, resulting in other diseases such as left displacement of abomasum and subclinical ketosis (LeBlanc et al., 2005). 

Dystocia, uterine prolapse and retained placenta 

Cows with clinical and subclinical hypocalcemia are at increased risk of dystocia, retained placenta and metritis. The loss of muscle tone in the uterus due to hypocalcemia increases the incidence of dystocia, uterine prolapse and retained placenta. It has been reported that milk fever affected cows are up to three times more likely to develop dystocia. In some cases the increased odds of dystocia were reported as six times in hypocalcemic cows than that of normal ones. Furthermore, dystocia can increase the risk of occurrence of retained placenta. The association of retained placenta and hypocalcemia has been reported. Erb et al. (1985) determined that cows with hypocalcemia were two times as likely to have retained placenta. Another study showed that cows with retained placenta had lower plasma concentrations of Ca at parturition and up to 7 days after parturition than cows without retained placenta. In addition, it has been reported that cows suffering from uterine prolapse have a lower serum Ca concentration than normal cows. It is worthy to bear in mind that retained placenta is a multi-etiological condition with many risk factors. Therefore, any association does not mean causal relationship. On the other hand, the pathogenic process leading to retained placenta is initiated before parturition (Melendez et al., 2004) and the serum level of Ca, after parturition cannot be considered a risk factor for retained placenta. 

Metritis and endometritis 

Subclinical hypocalcemia has been related to metritis. Because under hypocalcemic conditions, immune function may be impaired and muscle contraction diminished, metritis is more prone to occur. Martinez et al. (2011) studied 110 cows in one herd in Florida, and demonstrated that cows with Ca <2.14 mmol/L at least once between 0 and 3 days in milk had 4.5-fold increased odds of me- tritis. In a recent study, multiparous cows with subclinical hypocalcemia had 4.85 greater odds of having metritis compared with normocalcemic multiparous cows. A significantly higher in- cidence rate of endometritis was observed in UK cows that suffered clinical hypocalcemia in com- parison to normocalcemic cows (Whiteford and Sheldon, 2005). Mastitis It was reported that cows with clinical milk fever were eight times more likely to develop mastitis than normal cows. Hypocal- cemia reduces teat sphincter contraction, thus, an open teat canal invites environmental pathogens to enter the mammary gland. On the other hand, hypocalcemic cows tend to spend more time ly- ing down than do normocalcemic animals, which could increase teat end exposure to environmental opportunist organisms. In addition, hypocalcemia has deleterious effect on peripheral I blood mononuclear cells function and this exacer- bates periparturient immunosuppression (Kimura et al., 2006).

Reproduction performance 

The association of clinical hypocalcemia and decreased fertility was reported in several studies. Some reports showed that there are no differences in the incidence of uterine diseases, services per conception, or days open when comparing hypocalcemic cows with normocalcemic ones. However, there are plentiful evidence indicated that hypocalcemia may cause infertility. A UK study reported an increased number of services per conception, an increased calving to first service interval and an increased calving to conception interval for clinical hypocalcemic dairy cows. Martinez et al. (2012) found that pregnancy rate and interval between calving and pregnancy were reduced under hypocalcemia. It has been suggested that clinical hypocalcemia results in reduced fertility in dairy cows due to its effect on uterine muscle function, slower uterine involution and reduced blood flow to the ovaries. Cows with clinical hypocalcemia had a greater diameter of the gravid uterine horn and non-gravid uterine horn between 15 and 45 days post-partum (indicative of slower uterine involution) and a significantly reduced likelihood of having a corpus luteum (indicative of ovulation since parturition) than normal cows. Furthermore, subclinical hypocalcemia affected reproductive performance such as estrous cyclicity and pregnancy rate to first AI. The odds of expressing estrus before 60 days in milk were lower in subclinical hypocalcemic cows than in normocalcemic ones (Whiteford and Sheldon, 2005). Caixeta et al. (2017) reported that cows with normocalcemia were 1.8 times more likely to return in estrus by the end of the voluntarily waiting period than cows classified as having subclinical hypocalcemia. Higher Ca concentrations from week -1 through week 3 relative to calving were associated with increased odds of pregnancy. The odds of conceiving was 1.5 times higher for cows with pre-calving Ca >2.3 mmol/L, and 1.3 times higher for cows with Ca >2.2 mmol/L in week 1, >2.3 mmol/L in week 2, and >2.4 mmol/L in week 3 relative to calving. In addition, it was reported that subclinically hypocalcemic cows have fewer ovulatory sized follicles at days 15, 30 and 45 post-partum and smaller follicles at first ovulation than normal cows. It should be emphasized that trying to improve fertility in dairy herds without first having an appropriate hypocalcemia prevention strategy will bring only limited improvements (Seifi and Kia, 2017).

6. Clinical signs  

Hypocalcaemia can be clinical or subclinical based on whether an animal may or may not show clinical signs. Clinical milk fever is the most severe hypocalcaemia results in a cow that is unable to rise (from lying to standing position) and is the most easily recognized form of hypocalcaemia with blood calcium concentration less than 5 mg/dL (Goff, 2008a). Subclinical hypocalcaemia results in less severe disturbances in blood Ca and does not have any outward sign. During subclinical hypocalcaemia, blood calcium concentration ranges between 5.5 and 8.0 mg/dL (Fikadu et al., 2016). 

The symptoms include, initially the animal is ataxic, nervous and hyperactive. There is poor appetite, reduced rumen motility, bloating, Low body temperature, slow respiration, impalpable pulse, weak but rapid heart beats (80-100 per minutes) with very hard to hear due to reduced ability of muscles to contract, dilated pupils and dry muzzle are a common signs. Other symptoms include turned head back to the flank, splayed out hind legs, paresis (difficulty to rise from lying down). Finally, coma and sudden death may occur (Oetzel, 2022). 

Based on the degree of hypocalcaemia and time of occurrence, the clinical signs of milk fever divided into three stages. Stage I milk fever is early signs without recumbency. It may go unnoticed because its signs are subtle and transient. Affected cattle may appear excitable, nervous, or weak. Cows in stage II milk fever are in sternal recumbency. They exhibit moderate to severe depression, partial paralysis and typically lie with their head turned into their flank (Oetzel, 2022). Body temperature is subnormal, muzzle dry and the heart rate will be rapid. Stage III hypocalcemic cows are completely paralyzed, typically bloated, in lateral recumbency and progressively loss consciousness that leading to coma. There is a marked fall in temperature and increased heart rate. Cows will not survive more if not treated (Engdawork, 2019).

7. Diagnosis

Diagnosis of milk fever is based on history taking, clinical examination and laboratory diagnosis. During history taking all the detailed information of the cow including age, breed, stage of lactation, milk yield and calving day should be collected. Milk fever commonly occurs in mature dairy cows usually 5-9 years old, within 72 hours after parturition. The incidence increases in high producing dairy cows. Jersey breeds are commonly affected cows. The laboratory determination of blood calcium level and good response to intravenous calcium solutions are the most accurate method to diagnose a case of milk fever. The normal serum Ca concentration is 8 - 10 mg/dL (Thirunavukkarasu et al., 2010). Cows with serum calcium lower than 7.5 mg/dL are as considered as hypocalcaemic. Animals with serum calcium level of 5.5 to 7.5 mg/dL show sign of stage I hypocalcaemia. Stage II hypocalcaemia seen with calcium levels of 3.5 to 6.5 mg/dL and stage III seen when calcium concentration falls below 3.0 mg/dL. Blood samples are often taken later if there has been no improvement (Allgrove, 2003).

Differential diagnosis

The differential diagnosis of milk fever can be subdivided as metabolic, toxemia and traumatic disorders. Metabolic diseases showing symptoms resembling milk fever include hypoglycemia, hypomagnesaemia and hypophosphotamia. Toxic conditions that create doubt with milk fever include acute toxic mastitis and acute diffuse peritonitis. Traumatic conditions may cause symptoms resembling milk fever includes maternal obstetrical paralysis and musculoskeletal injury including downer cow’s syndrome due to pressure damage to muscles and nerves. Most cases can be differentiated from milk fever, as hypocalcaemia has a rapid response and good recovery to administration of IV calcium borogluconate (Bewley and Schutz, 2008).

8. Treatment

Treatment of milk fever should be done as early as possible, especially if recumbency is present, as recumbency can cause severe musculoskeletal damage. Commonly milk fever is treated with oral calcium solutions and intravenous (IV) calcium borogluconate. Supplementation of calcium borogluconate by oral route is the best approach to hypocalcemic cows that are still standing, such as cows in stage I hypocalcemia or which have undetected subclinical hypocalcemia. Cows absorb an effective amount of calcium into its bloodstream within about 30 minutes of supplementation. Drenching of calcium borogluconate near calving also serves as the prevention of milk fever (Goff, 2004). The fastest way to restore normal plasma calcium concentration is to administer an IV injection of calcium salts. For cows in stage II and III of milk fever should be treated immediately with a slow IV administration of 500 ml of 23% calcium borogluconate. Extremely high dose of calcium may cause fatal cardiac complications. Subcutaneous calcium administration can also be used to support blood calcium concentrations around calving. Subcutaneous calcium injections are irritating causes tissue necrosis; administration should be limited to no more than 75 ml of a 23% calcium borogluconate. The prognosis depends on the stage of the condition; cows in severe stage may present several complications and poor prognosis (Hutjens and Aalseth, 2005).

9. Control and prevention

Herd-based tests are now available for use in routine herd monitoring and for investigating dairy herds with metabolic subclinical problems. It has been suggested that 12 multiparous cows to be sampled within 48 hour after calving. Moreover, the results to be interpreted as the proportion of cows below the cut points of Ca. Besides defining the appropriate cut points for tests evaluated as proportion outcomes, it is also necessary to determine the alarm level for the proportion of animals below the described cut point. Because of normal biologic variation, a few individual cows are expected to be below the biologic threshold in any dairy (Oetzel, 2004).

9.1 Ca cut-off levels 

Cows with a serum Ca concentration less than 2.0 mmol/L were considered as hypocalcemic. Either pre- or post-partum cows with serum total Ca below 2.0 mmol/L were four times more likely to have post-partum disease problems . Although this is a conservative threshold, it is well accepted in research and clinical practice (Wilhelm et al., 2017). 

Recently higher thresholds were associated with a negative health outcome such as displacement of abomasum and metritis or an increased culling risk. These associations, however, found in a longer risk period before or after calving. Neves et al. (2017) has shown that prepartum cows with Ca concentrations metritis compared with normocalcemic cows (Martinez et al., 2012). Furthermore, Van Saun (2000) considered serum Ca concentration below 2.25 mmol/L as a cut-off level for interpretation of hypocalcemia in a group of fresh cows whereas it would be considered normal in an individual. Further evaluations are necessary to define the most appropriate threshold of hypocalcemia within any given time during transition period (Venjakob et al., 2017). 

9.2 Interpretation of the herd based Ca testing 

Because the duration of parturient hypocalcemia is extremely short (about the first 48 hours after calving), its incidence is monitored instead of its prevalence. Limited data are available to assist in determining an alarm level for parturient hypocalcemia. In regard to incidence of hypocalcemia, it was suggested that 30% hypocalcemia is a reasonable alarm level in multiparous Holstein cows (Oetzel, 2004). 

In authors’ experience, the best time to collect blood samples is about 12 to 24 hours. Some researchers suggested cows to be sampled within 48 hours after calving. Further studies are needed to determine the best sampling time and the accurate and precise cut-off level for either sampling time. 

Herds were categorized based on the proportion of positive samples (i.e., blood Ca below threshold) into negative (0 to 2 out of 12 cows), borderline (3 to 5 out of 12 cows), or positive (= 6 cows out of 12). Such classification is based on the assumptions provided by Oetzel (2004) using a 75% confidence interval and an alarm level of 30%. The cows sampled within 48 hour after parturition and serum Ca below 2.0 mmol/L were considered as hypocalcemic. It is needed to emphasize that this alarm level is considered to predict clinical hypocalcemia. Therefore, for predicting subclinical hypocalcemia, the alarm level may be lower. Some herds may be classified as borderline. In such cases, Venjakob et al. (2017) advise to draw more samples to classify the herd more appropriately (Venjakob et al., 2017).

 10. References 

Allgrove, J., 2003. Disorders of calcium metabolism. Current Paediatrics 13, 529-535.

Bewley, J., Schutz, M., 2008. An interdisciplinary review of body condition scoring for dairy cattle. The professional animal scientist 24, 507-529.

Chapinal, N., Carson, M., Duffield, T., Capel, M., Godden, S., Overton, M., Santos, J., LeBlanc, S., 2011. The association of serum metabolites with clinical disease during the transition period. Journal of dairy science 94, 4897-4903.

Constable, P.D., Hinchcliff, K.W., Done, S.H., Grünberg, W., 2016. Veterinary medicine: a textbook of the diseases of cattle, horses, sheep, pigs and goats. Elsevier Health Sciences.

Engdawork, A., 2019. Milk fever and its economical impacts in commercial dairy cattle production.

Fikadu, W., Tegegne, D., Abdela, N., Ahmed, W.M., 2016. Milk fever and its economic consequences in dairy cows: a review. Global veterinaria 16, 441-452.

Gallo, E.M., Canté-Barrett, K., Crabtree, G.R., 2006. Lymphocyte calcium signaling from membrane to nucleus. Nature immunology 7, 25-32.

Goff, J.P., 2004. Macromineral disorders of the transition cow. Veterinary Clinics: Food Animal Practice 20, 471-494.

Goff, J.P., 2008a. The monitoring, prevention, and treatment of milk fever and subclinical hypocalcemia in dairy cows. The veterinary journal 176, 50-57.

Goff, J.P., 2008b. Transition cow immune function and interaction with metabolic diseases, Tri-state dairy nutrition conference. Citeseer, pp. 45-57.

Goff, J.P., 2014. Calcium and magnesium disorders. Veterinary Clinics: Food Animal Practice 30, 359-381.

Hamali, H., 2008. Post estrus hypocalcemia in a repeat breeder halfbreed Holstein cow. J. Anim. Vet. Adv 7, 1301-1304.

Hanai, H., Brennan, D., Cheng, L., Goldman, M., Chorev, M., Levine, M., Sacktor, B., Liang, C., 1990. Downregulation of parathyroid hormone receptors in renal membranes from aged rats. American Journal of Physiology-Renal Physiology 259, F444-F450.

Holler, H., Breeves, G., Kocabatmax, M., Gredes, H., 1988. Mineral absorption across isolated rumen mucosa. Queensland Journal of Experimental Physiology 73, 609-611.

Hutjens, M., Aalseth, E.P., 2005. Caring for transition cows. Hoard's Dairyman Books.

Kimura, K., Reinhardt, T., Goff, J., 2006. Parturition and hypocalcemia blunts calcium signals in immune cells of dairy cattle. Journal of dairy science 89, 2588-2595.

LeBlanc, S., Leslie, K., Duffield, T., 2005. Metabolic predictors of displaced abomasum in dairy cattle. Journal of dairy science 88, 159-170.

Martín-Tereso, J., Martens, H., 2014. Calcium and magnesium physiology and nutrition in relation to the prevention of milk fever and tetany (dietary management of macrominerals in preventing disease). Veterinary Clinics: Food Animal Practice 30, 643-670.

Martinez, N., Risco, C., Lima, F., Bisinotto, R., Greco, L., Ribeiro, E., Maunsell, F., Galvão, K., Santos, J., 2012. Evaluation of peripartal calcium status, energetic profile, and neutrophil function in dairy cows at low or high risk of developing uterine disease. Journal of dairy science 95, 7158-7172.

Martinez, N., Sinedino, L., Bisinotto, R., Ribeiro, E., Gomes, G., Lima, F., Greco, L., Risco, C., Galvão, K., Taylor-Rodriguez, D., 2014. Effect of induced subclinical hypocalcemia on physiological responses and neutrophil function in dairy cows. Journal of dairy science 97, 874-887.

Melendez, P., Donovan, G.A., Risco, C.A., Goff, J.P., 2004. Plasma mineral and energy metabolite concentrations in dairy cows fed an anionic prepartum diet that did or did not have retained fetal membranes after parturition. American Journal of Veterinary Research 65, 1071-1076.

Mulligan, F., Doherty, M., 2008. Production diseases of the transition cow. The Veterinary Journal 176, 3-9.

Oetzel, G., 2022. Non-infectious diseases: Milk fever.

Oetzel, G.R., 2004. Monitoring and testing dairy herds for metabolic disease. Veterinary Clinics: Food Animal Practice 20, 651-674.

Reinhardt, T.A., Lippolis, J.D., McCluskey, B.J., Goff, J.P., Horst, R.L., 2011. Prevalence of subclinical hypocalcemia in dairy herds. The Veterinary Journal 188, 122-124.

Rodríguez, E., Arís, A., Bach, A., 2017. Associations between subclinical hypocalcemia and postparturient diseases in dairy cows. Journal of dairy science 100, 7427-7434.

Schnewille, J.T., van ‘T Klooster, A.T., Beynen, A., 1994. High phosphorus intake depresses apparent magnesium absorption in pregnant heifers. Journal of Animal Physiology and Animal Nutrition 71, 15-21.

Seifi, H.A., Kia, S., 2017. Subclinical hypocalcemia in dairy cows: Pathophysiology, consequences and monitoring. Iranian Journal of Veterinary Science and Technology 9, 1-15.

Seifi, H.A., Mohri, M., Ehsani, A., Hosseini, E., Chamsaz, M., 2005. Interpretation of bovine serum total calcium: effects of adjustment for albumin and total protein. Comparative Clinical Pathology 14, 155-159.

Seifi, H.A., Mohri, M., Zadeh, J.K., 2004. Use of pre-partum urine pH to predict the risk of milk fever in dairy cows. The veterinary journal 167, 281-285.

Thirunavukkarasu, M., Kathiravan, G., Kalaikannan, A., Jebarani, W., 2010. Quantifying economic losses due to milk fever in dairy farms. Agricultural Economics Research Review 23, 77-82.

Thrall, M.A., Weiser, G., Allison, R.W., Campbell, T.W., 2012. Veterinary hematology and clinical chemistry. John Wiley & Sons.

USDA, 2016. Dairy 2014: Dairy cattle management practices in the United States, 2014. USDA–APHIS–VS–CEAH–NAHMS. Fort Collins, CO# 692.0216.

Venjakob, P., Borchardt, S., Heuwieser, W., 2017. Hypocalcemia—Cow-level prevalence and preventive strategies in German dairy herds. Journal of dairy science 100, 9258-9266.

Whitaker, D., 2000. Use and interpretation of metabolic profiles. The health of dairy cattle, 89-107.

Whiteford, L., Sheldon, I., 2005. Association between clinical hypocalcaemia and postpartum endometritis. The Veterinary Record 157, 202.

Wilhelm, A., Maquivar, M., Bas, S., Brick, T., Weiss, W., Bothe, H., Velez, J., Schuenemann, G., 2017. Effect of serum calcium status at calving on survival, health, and performance of postpartum Holstein cows and calves under certified organic management. Journal of dairy science 100, 3059-3067.