Evidence for and against manganese deficiency as causal for congenital joint deficiency disease or death in fetal and neonatal cattle

Abstract

Measures of manganese (Mn) status in cattle vary among studies, and no single criterion accurately predicts or diagnoses Mn deficiency and pathologic outcomes. Mn deficiency causes congenital joint laxity and dwarfism (CJLD) when total dietary intake is <20 ppm Mn dry matter (DM) for most of the pregnancy. However, the recommended dietary intake of 40 ppm DM can also result in clinical Mn deficiency. Some studies have found that CJLD occurs in calves from cows fed red clover or silage but not in calves from cows fed hay. The concentration of Mn in the liver is the best indicator of Mn status in neonates and adults but cannot be interpreted in fetuses. Serum, plasma, and whole blood concentrations of Mn are unreliable indicators of bovine Mn status. The primary objective of our report is to present evidence linking CJLD to a primary or secondary Mn deficiency. To predict and diagnose Mn deficiency in cattle, we propose using a combination of clinical signs, dietary Mn, liver Mn at birth and beyond, positive response to Mn supplementation or the replacement of silage with other forages, and ruling out other causes of malformations. By following these recommendations, we expect that CJLD and gestational death will decrease as hepatic Mn concentrations increase at birth. Many publications we reviewed are not statistically sound, and future research should include a statistician from the initial discussions of the study through the final publication.

Bone is the tissue that is most responsive to dietary intake of manganese (Mn) because Mn is essential for the formation of chondroitin sulfate as a component of glycosaminoglycans in the organic bone matrix. ¹³ The role of Mn in cartilage formation makes it essential to the formation of the epiphyseal growth plate, which directly affects longitudinal bone growth. The most frequent signs of Mn deficiency in young animals are skeletal malformations, in particular, congenital joint laxity and dwarfism (CJLD),^{1,2,12,18,20,24} namely, twisted and short limbs, swollen joints, disproportionate dwarfism, superior brachygnathism, and shortened nasomaxillary bones in which the short upper jaw exposes the mandibular incisors. Although Mn was one of the earliest trace minerals to be recognized as nutritionally essential, ¹ limited scientific research has made it one of the least understood vital metals. Here, we discuss the interpretation of different measures used to diagnose Mn deficiency in cattle.

No current criterion predicts Mn deficiency in all cattle. There is a lack of robust control studies that consistently prevent gestational bone malformation and death over the range of dam and fetal ages. This lack of data affects the reliability of tissue and feed Mn concentrations to ensure production herd health. ³ Notwithstanding this issue, the Mn concentration in feedstuffs is considered the best indicator of dietary adequacy. ¹⁷ This belief is correct for low dietary Mn concentrations (≤20 ppm dry matter [DM]), although offspring with CJLD can be produced at the National Research Council (NRC)-recommended intake of 40 ppm DM.^10,16

To evaluate evidence that Mn deficiency is causal for CJLD, we reviewed all accessible papers and theses identified by the CABI digital library, EBSCO Discovery Service, Google Scholar, ProQuest Digital Dissertations, PubMed, SciFinder, Universidad de Antioquia and the University of Illinois Library online collections, and WorldCat Collection based on the keywords “acorn calves,” “beef, calves, cattle,” “chondrodysplasia”, “CJLD”, “congenital abnormality”, “dairy”, “death”, “Bulldog calves diet”, “gestation”, “liver”, “manganese (Mn)”, “Mn absorption”, “Mn metabolism”, “Mn tissue concentration”. Here, we summarize the evidence for and against the pivotal role of Mn in CJLD.

Mn requirements for dairy cattle have changed in various editions of the NRC guidelines; the 1984 6th edition NRC guidelines of 40 mg of Mn/kg of DM for all classes of cattle were lowered to 17.8 ppm Mn of DM in the 2001 7th edition, ¹⁶ and unchanged in the 2016 8th edition. ¹⁷ This concentration is similar to Mn concentrations that produce deficiency signs in beef cows. When apparent digestibility and retention of Mn were later performed in lactating and dry dairy cows, the requirements were 1.6 and 2.7 times higher than concentrations calculated using the 2001 7th edition NRC model. ²³ Based on these recommendations, the level of dietary Mn below which Mn stores and Mn-dependent processes in the body begin to be affected is unclear and should be further refined. The effects of Mn deficiency on anestrus and infertility are likely to occur at ration Mn concentrations lower than those affecting growth performance or causing CJLD. ¹ Compared to 30 ppm Mn DM in heifer diets, heifers fed 7–10 ppm Mn DM had delayed onset of first estrus, a slightly reduced conception rate, and more calves born with weak legs and pasterns at calving. ¹ Optimal bovine embryo development to the blastocyst stage partially depends on Mn presence during in vivo maturation. ¹³ Mn might create suboptimal progesterone secretion during pregnancy days 4–11, which results in early embryonic loss. ¹³

CJLD was reproduced when Mn concentrations were ≥40 ppm DM ¹⁰ in beef cows wintered on red clover or timothy grass silage but not when fed timothy hay harvested from the same fields. ¹⁰ The incidence of CJLD in calves born to cows fed red clover (8 of 21; 38%) or timothy grass silage (14 of 52; 28%) did not occur in calves born to cows fed timothy hay (0 of 24; 0%). The average Mn concentration in the hay (51 ppm DM) was lower than in the silages (64 or 63 ppm DM). Other field case reports also linked CJLD to feeding clover or grass silage but not hay harvested from the same fields.^2,15,18 Affected calves also had an increased peripartum mortality rate, and many of those that lived did not reach normal stature as yearlings. ¹⁸ Although dietary Mn was not always measured, these studies reported that supplementing the silage with hay eliminated the problem; grain reduced the risk to a lesser extent.^2,15,18 For example, losses of 2–46% of the calf crop in consecutive years were eliminated when the grass/clover silage diet was supplemented with 2.5–4.5 kg/head/d of hay and 0.75 to 1.5 kg/head/d of rolled barley and was more effective than without hay. ¹⁸ Some authors proposed Mn involvement in CJLD but did not demonstrate that Mn deficiency was definitively causal. They also suggested that a process during silage storage may transform the grass into a teratogenic feed when fed to pregnant cows as the exclusive diet.

Soil contamination of harvested feeds (e.g., corn greenchop) before ensiling can be a significant source of bioavailable dietary iron (Fe) that can negatively affect Mn absorption. ⁹ Soil Fe is often in the ferric form bound in insoluble complexes; however, exposure to an acidic environment similar to that occurring during silage fermentation can facilitate reduction to the more soluble ferrous form. ⁹ Feeding calves a diet high in soluble Fe (as FeSO₄) decreased the concentrations of divalent metal transporter 1, which transports Fe and Mn. ⁶ Liver Fe increased significantly from 117 ppm DM in controls to 153 ppm DM (~30 and 40 ppm wet weight [WW]) in the high-Fe diet group, but liver concentrations of Mn were not affected. ⁶

Chondrodystrophic calves with concurrent congenital goiter have been ascribed to the secondary antagonistic interaction of Fe with Mn. ¹² Cattle with CJLD that had been fed silage might also have been affected by Fe antagonism of Mn availability.^5,10,15,18 Based on the changes resulting from diet differences, several cited studies reported that a cause of CJLD is a Mn deficiency due to its low bioavailability in clover or grass silage.^6,7,9,10 Specific recommendations for affected herds include replacing “a quarter of the silage dry matter intake with alternative fodder (e.g., hay or straw), and/or barley [as] pulp or meal.” ¹⁵ Feed analysis provides an accessible estimate of dietary Mn intake for dairy cattle but not beef cattle because consumption is unregulated and plant Mn concentrations vary across fields. The feed concentrations alone do not estimate bioavailability.^16,23 CJLD can be expected when Mn concentrations in the diet are <20 ppm DM throughout most of the pregnancy^7,8,16 but not at 40 ppm DM. ¹⁶ Authors of a later study ¹¹ concluded that “the high plasma phosphorus concentrations in CJLD-affected calves and their dams could be related to the aetiology of the CJLD condition in calves [because] phosphorus interferes with manganese absorption.” Only cows fed grass silage but not hay from October 15 to calving in March gave birth to calves affected by CJLD, ¹¹ but Mn was not measured. Some field outbreaks have associated high circulating phosphorus concentrations with Mn below the RI in CJLD-affected calves ² ; CJLD did not recur after alternative forages replaced home-produced grass silage. ²

There is no evidence that plasma, serum, or whole blood Mn concentrations are reliable indicators of Mn biologic status. Tests of tissue concentrations have been highly variable among studies, and no criteria have been demonstrated to predict Mn deprivation reliably. The suggested whole blood cutoff value for Mn deficiency is 0.02 ppm. ¹⁷ Whole blood concentrations ≥0.02 Mn ppm have been detected at birth in calves with CJLD from heifers unsupplemented starting 4 mo before conception.^7,8 In those studies, the addition of 50 ppm of Mn to the control (deficient) diet (15.8 ppm DM) did not affect the whole-blood Mn concentration of heifers, and the calves born with CJLD also had blood concentrations >0.02 ppm (x- of 0.24 vs. 0.35 ppm in calves from deficient and supplemented heifers, respectively). These studies^7,8 and others ¹ support that a diet containing <20 ppm DM is insufficient for proper fetal development in gestating heifers despite not affecting their whole-blood concentrations. Ten control heifers and 10 heifers supplemented with 50 ppm DM remained on study to day 463. ⁸ Similar whole-blood Mn concentrations on day 463 for these 2 groups indicated a rigorous homeostatic mechanism for Mn. ⁸

The exact time frame in which Mn deficiency results in cases of CJLD has not been elucidated. Experimental studies did not reproduce cases of CJLD when a diet was low in Mn starting at the time of breeding in ~2-y-old heifers, although calves were born weak and had difficulty standing compared to calves from heifers fed an adequate diet. ¹⁴ However, 5 feeding trials feeding silage on 3 Canadian ranches with cases of CJLD showed that the gestation period during which the fetus is susceptible to CJLD spans from at least day 107 to day 230. ¹⁸ Therefore, it was recommended that silage be supplemented with hay and grain from the start of overwinter feeding and that this supplementation be continued until calving. The reports also describe other skeletal defects, such as crooked calf disease, but none matched the description of CJLD. A 2019 survey of the mineral status of California beef cattle in 50 herds (555 animals) ³ found that 92% of cattle sera fell below the 5-ppb guideline for Mn, ¹⁷ but did not report disease. It is unlikely that such a large number of tested animals were truly deficient. An experimental study with pregnant cows fed hay, red clover, silage, or grass silage throughout gestation reported serum Mn concentrations of 2–3 ppb in all groups. ¹⁰ However, only the offspring of the groups fed red clover (38%) or grass (28%) silage developed CJLD. In 2 other studies that measured plasma or serum Mn concentrations, no difference was found in steers and heifers fed graded concentrations of Mn.^7,8 The authors concluded that the consistently low concentrations of Mn in plasma, regardless of dietary Mn concentration, implied an effective mechanism for Mn homeostasis.

The liver concentration of Mn indicates the body Mn status accurately in neonates and adults but not in fetuses. Fetal and calf liver and kidney concentrations may be low at <1 ppm WW, reaching normal concentrations by 2–3-wk-old. ¹⁷ However, in field and experimental studies of newborn calves with CJLD, the liver concentrations were 1.0–2.0 ppm WW or lower.^12,14 Manganese concentrations in normal calves at birth were ≥2 ppm WW. However, calves that appear normal can have liver Mn concentrations of <2 ppm WW at birth, which is a problem with attributing CJLD to Mn deficiency; if 1–2 ppm is too low, then most such calves should, but do not, exhibit the syndrome. Therefore, other minerals (e.g., Fe, P), individual metabolic profiles, genetics, and other factors likely moderate the developmental effects of Mn.

The growth, development, and reproductive performance of beef heifers individually fed graded concentrations of Mn (0, 10, 30, or 50 ppm DM) from 10–16 mo of age were recorded during a 196-d trial. ⁷ Starting at a basal diet of 15.8 ppm DM (control), liver biopsy samples on days 98 and 196 had mean Mn concentrations of 2.25–2.75 ppm WW in all groups.^7,8 No adverse effects were observed in growth characteristics and reproductive performance (pregnancy rate, conception rate, age at conception, and services to conception) in the unsupplemented control and supplemented groups. At termination, 10 pregnant heifers from the control group (15.8 ppm DM) and 10 from the 50 ppm group remained on their respective diets throughout gestation and early lactation. ⁸ All calves born to the controls, but not the supplemented group, showed some or all typical signs of CJLD. Although the initial body stores provided adequate Mn during the 196-d before gestation, ⁷ stores were insufficient to support fetal development throughout the gestation period. ⁸ The liver Mn concentration was not measured in calves born with CJLD.

Liver Mn concentrations in bovine fetuses and their corresponding dams were collected during routine slaughter operations in 2 studies,^4,5 but neither reported Mn in feedstuffs. One study found no differences in liver Mn concentrations in different classes of adults (pregnant, nonpregnant, heifers, steers). ⁴ Mean liver Mn concentrations in dams were 2.1–2.3 ppm WW and ~60–80% in their fetuses (1.4–1.6 ppm WW). However, there was a large standard error for concentrations in each trimester. The second study considered the “cattle to be representative of slaughtered Holstein dairy cattle in California.” Apart from correlating maternal and unaborted fetal hepatic Mn concentrations at slaughter, “normal” size-matched unaborted fetuses (n = 103) were compared to samples submitted by veterinarians for routine diagnosis of aborted fetuses (n = 80). ⁵ The mean maternal liver Mn concentrations in each trimester were 3.5, 3.2, and 3.1 ppm, and fetal concentrations were 1.7, 1.7, and 2.1 ppm. The ranges in each trimester for maternal, unaborted, and aborted fetuses were too large to estimate normal values. However, the lower liver concentrations of Mn (and Cu, Fe, Zn) in aborted than in unaborted fetuses “suggest a nonspecific change in trace element status, which implies an effect of abortion, not a cause of abortion.” ⁵ Liver Mn concentrations were consistently higher in fetuses as the stage of pregnancy approached parturition. ⁵

Similar findings from 501 beef calves and fetuses in western Canada at postmortem examinations were classified into abortion, stillbirth, neonatal, and postnatal to assess the age effect. ²² These authors proposed empirical prediction intervals for each age group and compared them with average Mn concentrations in liver. ²² No skeletal deformities were reported in any of the calves analyzed for Mn. At 3 farms over consecutive years, the x̅ ± SD liver Mn concentrations (WW) for all calves were 1.88 ± 5.08 (n = 225), 1.30 ± 0.16 (n = 80), and 1.38 ± 3.42 ppm (n = 196). ²² At the 3 farms, liver Mn was higher in neonatal (1.95 ± 0.34, 1.49 ± 0.15, no record) and postnatal calves (>3 d to <3 mo) that died (2.24 ± 5.14, 2.20 ± 0.13 ppm, no record) than calves dead from either abortion (1.34 ± 0.07, 1.19 ± 0.18, 0.86 ± 0.35 ppm) or stillbirths (1.20 ± 6.47, 0.90 ± 0.15, 0.76 ± 0.11 ppm). ²² The rate of death, defined as the slope of the liver Mn concentration (WW) to the percentile of dead calves, increased with both the age at death and the liver Mn concentration

These studies highlight the difficulties in linking tissue Mn concentrations with abortion or CJLD. Some calves with CJLD had liver Mn concentrations ≤2 ppm WW ¹² ; however, most calves with liver Mn ≤2 ppm WW had no signs of chondrodysplasia or dwarfism.^14,21 Because some control calves have Mn concentrations <2 ppm, clinical defects are one extreme with subclinical diseases at the other end of the deficiency-associated disease spectrum. ²⁵

In conclusion, a single tissue measurement cannot determine Mn deficiency as the cause of a pregnancy or congenital disorder. Mn deficiency in CJLD, which is the presumed classic presentation, should be supported by low fetal liver Mn, the type of feed during gestation (i.e., silage), high serum Ca and P concentrations and ratios in dams and affected calves, positive response to Mn supplementation, or change of a silage diet for hay and grains, in the following gestation. Dietary Mn should be >20 ppm DM, except that when silage is fed, the requirements could increase due to a secondary deficiency caused by Fe, P, and Ca antagonism.^4,5,6,21 Reproductive problems can occur on lower Mn diets that cause CJLD (typically ≤10 ppm DM). Serum and whole blood concentrations of Mn in affected newborn calves could be within the RI but should not be considered definitive for normalcy. Similarly, dams could have normal blood and serum Mn concentrations and still be deficient. From the literature, Mn deficiency is suspected in CJLD cases if: 1) congenital malformations fit with classical CJLD ascribed to Mn deficiency; 2) dams were fed a deficient diet rather than a diet that prevented absorption, such as silage, for most of the pregnancy^10,11,14; 3) although liver concentrations in most normal animals are >2 ppm, hepatic Mn ≤2 ppm would be suggestive of Mn deficiency, and 4) there was a positive response to Mn supplementation in the subsequent pregnancy.

New studies should involve a statistician from the conception to the conclusion and publication. ¹⁹ The study design and analysis should follow international protocols for quantifying Mn in feeds, dams, fetuses, and neonates. Even then, “mineral balances are difficult to interpret because of the excretion of the respective elements in feces. Attempts to [mathematically analyze] this situation are not very successful because modulation of absorption leads to large variation in estimates of true availability.” ²¹