The assessment of blood copper status in cattle: a comparison of measurements of caeruloplasmin and elemental copper in serum and plasma

AIM: To evaluate the effect of test, either copper (Cu) concentration or caeruloplasmin (CP) activity, and sample type, either serum or plasma, on the diagnosis of blood Cu status in cattle. METHODS: Paired serum and heparinised plasma samples taken from 125 cattle in 13 herds were tested for Cu concentration and CP activity. The individual results for serum Cu concentration and serum and plasma CP activities were compared with the plasma Cu concentration results, as were their diagnostic values as determined by reference ranges, i.e. 'marginal', 'adequate', 'excess'. RESULTS: The overall mean serum Cu concentration was 2.92 micromol/L lower than the mean plasma Cu concentration; however, there was significant variability between individual samples, and the 95% limits of agreement ranged from 0.44 micromol/L more to 6.28 micromol/L less. The relationship between CP activity and plasma Cu concentration was less variable; the 95% prediction interval for plasma Cu concentration from CP activity was +/- 2.8 micromol/L, and was unaffected by whether CP activity was measured in plasma or serum. Using the threshold currently recommended for 'marginal' status of <8.0 micromol/L for serum Cu concentration identified a significantly different population of cattle than a threshold of <9.0 micromol/L for plasma samples. Altering the threshold to <7.0 micromol/L for serum Cu concentration produced better agreement. For CP activity, a threshold of 15 IU/L for both serum and plasma identified the same population as a threshold of <9 micromol/L for plasma Cu concentration. CONCLUSIONS: Serum Cu concentration is not a suitable substitute for plasma Cu concentration for the detection of 'marginal' blood Cu status in cattle as the individual variability in the apparent loss of Cu during clotting is too great. In this study, CP activity, in both serum and plasma, was found to be a suitable substitute for the detection of 'marginal' blood Cu status. CLINICAL RELEVANCE: The use of serum Cu concentration rather than plasma Cu concentration in the diagnosis of Cu responsive disease in cattle needs to be re-evaluated as does the way in which individual sample results are used in such tests.     

Comparison of concentrations of copper in plasma and serum from farmed red deer (Cervus elaphus)

AIM: To determine the relationship between the concentrations of Cu in plasma and serum in red deer, and to compare this relationship with those previously reported in cattle and sheep.

METHODS: Paired serum and heparinised plasma samples from 114 red deer from 10 herds (n=6-20 per herd) were analysed for concentrations of Cu. Samples were collected either at slaughter (n=84; eight herds) or by jugular venepuncture (n=30; two herds). Thirty-nine of the samples taken at slaughter were from adult hinds from four herds, while other samples were taken from 10–14-month-old males, except for one herd (10 samples) where an equal number of 8–9-month-old males and females were sampled. The effect of age, gender and herd on the relationship between concentrations of Cu in plasma and serum was assessed using univariate ANOVA. The individual results for concentrations of Cu in serum were compared with those in plasma, using limits-of-agreement plotting.

RESULTS: The mean concentration of Cu in plasma was not significantly different from that of serum (0.048; 95% CI=-0.14 to 0.24 µmol/L). There was no effect of age, sex or herd on this relationship.

CONCLUSIONS: In deer, there was no significant difference between concentrations of Cu in plasma and serum regardless of age, sex or herd of origin.

CLINICAL RELEVANCE: In contrast to the situation in cattle and sheep, the concentration of Cu in serum can be used interchangeably with that in plasma for the estimation of concentration of Cu in blood of red deer.     

Copper deficiency in sheep: An assessment of the relationship between concentrations of copper in serum and plasma

AIM: To assess the relationship between concentrations of copper in serum and plasma in sheep.

METHODS: Concentrations of Cu were measured in paired serum and heparinised plasma samples collected from 110 sheep in nine flocks. Linear regression was used to evaluate whether flock or gender had a significant effect on the association between concentrations of Cu in serum and plasma. The individual results for concentrations of Cu in serum were then compared with those from plasma, using correlation and limits of agreement plotting.

RESULTS: Concentrations of Cu in serum ranged from 7.3 to 22 (mean 14.0) µmol/L, while concentrations in plasma ranged from 9 to 27 (mean 16.3) µmol/L. On average, concentrations of Cu in serum were 2.3 µmol/L lower than in plasma. Over the range of values seen in this study, concentrations of Cu in plasma and serum were significantly correlated (r=0.89), and mean concentrations in serum were 87% of those in plasma. There was no effect of flock or gender on the relationship between concentrations of Cu in serum and plasma. Despite the significant correlation, there was marked variability between individual samples in the proportion of Cu that was lost during clotting, with the 95% limits of agreement for serum Cu ranging from 70 to 104% of the plasma concentration.

CONCLUSIONS: As in cattle, the individual variability in the loss of Cu during clotting in sheep is too great for concentration of Cu in serum to be used as a substitute for that in plasma.

CLINICAL RELEVANCE: When assessing the blood Cu pool as part of the diagnosis of Cu-responsive disease in sheep, the concentration of Cu in plasma should be measured in preference to that of serum. We suggest that a range of 4.5 to 9 µmol/L in plasma be used to define marginal Cu status in sheep.     

An evaluation of the copper sequestrated during clotting in cattle: is it just caeruloplasmin?

In cattle, sequestration of copper (Cu) occurs during the clotting process so that serum Cu concentrations are markedly lower than plasma Cu. It has been suggested that all of the Cu lost during clotting is caeruloplasmin (CP). This study used paired samples from 125 cattle to assess whether this assumption was correct. The regression equations for plasma CP activity against plasma Cu concentration and serum CP against serum Cu had significantly different intercepts suggesting that at zero CP activity the amount of Cu remaining was dependent on sample type. Furthermore, the difference between serum and plasma Cu was unrelated to Cu status, whereas the difference between serum and plasma CP was related to Cu status. The regression equation for the loss of CP activity against change in Cu concentration had an intercept that was different from zero, indicating that a reduction in Cu concentration could occur even if CP activity was unaffected by clotting.     

Impact of molybdenum on the copper status of red deer (Cervus elaphus)

AIM: To determine the effect of increasing molybdenum (Mo) intakes on serum and liver copper (Cu) concentrations and growth rates of grazing red deer (Cervus elaphus).

METHODS: Molybdenum- and Cu-amended fertilisers were applied to six 1.1-ha paddocks in a 3 × 2 design. Three levels of Mo were applied on two paddocks at each level in mid April (designated Day 1); levels were: none (control), 0.5 (medium) and 1.0 (high) kg Mo/ha as sodium molybdate. In late May (Day 39), two levels of Cu (none and 3.0 kg Cu/ha, as copper sulphate) were applied to each of the three levels of Mo-treated paddocks. Pasture Mo, Cu and sulphur (S) concentrations were measured at about fortnightly intervals. In late June (Day 74), ten 6-month-old red deer hinds were placed on the six experimental pastures, and serum and liver Cu concentrations were monitored at about monthly intervals for 102 days. The hinds were weighed on four occasions during the trial.

RESULTS: Mean pasture Mo concentrations on Day 56 were 2, 4.6 and 11.3 mg/kg dry matter (DM) for the untreated control, medium and high Mo-treated pastures, respectively. Pasture Cu concentration was 95 mg/kg DM on Day 59, 53 mg/kg DM on Day 90, and 9 mg/kg DM by Day 153. Mean S concentration in pasture was 3.3 (range 3.03–3.45) g/kg DM. Copper application to pasture had no significant effect on serum and liver Cu concentrations in deer so data were pooled within Mo treatment. Mean initial (Day 74) serum Cu concentration was 9.2 µmol/L. In the deer grazing the control Mo pasture, this increased to 10.3 µmol/L on Day 112, before decreasing to 6.4 µmol/L on Day 176. In deer grazing the medium and high Mo-treated pastures, mean serum Cu concentrations were 3.8 and 3.9 µmol/L, respectively, on Day 112, and 2.5 and 3.3 µmol/L, respectively, on Day 176. Mean initial (Day 74) liver Cu concentration was 131 µmol/kg fresh tissue. In the deer grazing the control Mo pasture, this declined to 120 and 52 µmol/kg on Days 112 and Day 176, respectively. In deer grazing the medium and high Motreated pastures, liver Cu concentrations decreased to 55 and 52 µmol/kg fresh tissue, respectively, on Day 112, and 21 and 20 µmol/kg fresh tissue, respectively, on Day 176. Mean serum and liver Cu concentrations were not significantly different between deer grazing the medium and high Mo-treated pastures, and were lower (serum p=0.003, liver p<0.001) in those groups than in deer grazing the untreated control pastures. No clinical signs of Cu deficiency associated with lameness were observed. Deer grazing pastures that had Mo concentrations >10 mg/kg DM had lower (p=0.002) growth rates (100 vs 130 g/day) than those on pastures containing <2.4 mg Mo/kg DM.

CONCLUSION: Increasing pasture Mo concentrations from 2 mg/kg DM to ≥4.6 mg/kg DM significantly reduced serum and liver Cu concentrations in grazing deer. Reduced growth rate was observed at pasture Mo concentrations >10 mg/kg DM.     

An evaluation of the effect of clotting on the relationship between copper and caeruloplasmin in bovine blood

The ratio of caeruloplasmin activity to copper concentration (CP:Cu) has been suggested as a more accurate determinant of the requirement for additional copper than plasma or liver copper concentrations. Although this test has no peer-reviewed evidence of efficacy, it has been used by a large number of UK veterinarians. However, the available test uses a serum caeruloplasmin (sCP) activity to plasma copper (pCu) concentration ratio which, because of the preferential loss of caeruloplasmin during clotting, is likely to underestimate the true CP:Cu, although it has been suggested that the marginal range accounts for this. This study was undertaken to evaluate the effect of using serum copper (sCu) rather than pCu concentrations in calculating CP:Cu. Using sCu rather than pCu increased CP:Cu by more than was accounted for by the marginal range. Of 48 cattle which were reported as 'low' using sCP:pCu, 22 were 'normal' when sCu was used instead of pCu. All herds with 'marginal' or 'low' mean CP:Cu when the sCP:pCu concentration ratio was used had 'normal' ratios when sCu was used instead of pCu.     

An evaluation of the effect of clotting on the recovery of copper from caprine blood

In the goat, diagnosis of copper (Cu) deficiency is often based on measurement of Cu in serum or plasma. Previous research in cattle and sheep has shown that these values are not interchangeable, but data for goats have not been published. Paired serum and heparinised plasma samples taken from 119 goats in eight herds were tested for Cu concentration. Plasma and serum Cu were significantly correlated (r=0.95). On average serum Cu was 3.5 μmol/L lower than plasma Cu, but this difference was related to Cu status (r=0.45). Mean serum Cu concentration was 83% of plasma Cu, with the 95% limits of agreement ranging from 66% to 100%. Similar to cattle and sheep, individual variability in Cu loss during clotting is too great for serum Cu to be used as a measure of Cu status in goats.     

Facial eczema management protocols used on dairy farms in the North Island of New Zealand and associated concentrations of zinc in serum

AIMS: To describe and evaluate the current practices used to manage and prevent facial eczema (FE) in North Island dairy herds, and determine the within-herd prevalence of cows with elevated activities of gamma glutamyl transferase (GGT), and with concentrations of Zn in serum <18?μmol/L.

METHODS: Between January and May 2014, 105 herd managers from throughout the North Island of New Zealand were invited to participate in the study when regional spore counts for Pithomyces chartarum started to rise towards 30,000 spores/g pasture. Managers selected 10 representative cattle that were weighed and blood-sampled by the herd veterinarian. Blood samples were analysed for concentrations of Zn in serum and GGT activity. Pasture samples were also collected and submitted for spore count estimation. Finally a survey of farm management practices relating to prevention of FE was completed by the herd manager. A mixed-effects logistic regression model was used to determine associations between herd-level and cow-level explanatory variables and the probability of a cow having a concentration of Zn in serum <18?µmol/L.

RESULTS: Of the 1,071 cows tested, 79 (7.3 (95% CI=5.8–9.0)%) had GGT activity in serum >300?IU/L, and 35/106 (33 (95% CI=24.2–42.8)%) herds had ≥1 of the 10 cows sampled with GGT activity >300?IU/L. Of the 911 cows that were being treated with Zn, concentrations of Zn were between 18–35?μmol/L in 398 (43.6 (95% CI=40.4–46.9)%) cows, were >35?μmol/L in 32 (3.5 (95% CI=2.4–4.1)%) cows, and <18?μmol/L in 479 (52.6 (95% CI=49.3–55.9)%) cows. After adjusting for the confounding effect of region, the odds of a cow having concentrations of Zn in serum <18?μmol/L were 5.5 (95% CI=1.1–29) times greater for cows supplemented with zinc in water compared with those supplemented by drenching. Of the 105 herd managers, 103 (98%) stated that they had access to regional spore count data, but only 35/105 (33%) reported that they measured spore counts on their own farm. Overall, 98/105 (93%) managers reported that they had some form of FE management programme in place. Fungicides were used on their own or in combination with zinc treatments in 10 herds, ZnSO₄ in water troughs was used in 68 herds, oral drenching with ZnO in nine herds, and ZnO supplied in-feed in 26 herds. Estimated daily dose rates of zinc were less than that required to treat a 400?kg cow on 42/68 farms that administered ZnSO₄ in the water or ZnO as a drench.

CONCLUSION AND CLINICAL RELEVANCE: This study has shown that FE management on dairy farms in the North Island of New Zealand could be substantially improved. It is likely that improved FE management would occur if herd managers were provided with more feedback on the success (or otherwise) of their FE management programmes.     

Skeletal myonecrosis in a cat

In this investigation electrical conductivity (EC) and somatic cell concentration (SCC) were compared in 3 herds for their ability to correctly identify the infection status of quarters. For EC, thresholds of 6.0 and 6.8 mS/cm were used and comparisons were made between quarters within each cow. The method of comparison between quarters was the same as that described by the manufacturers of the AHI Mastitis Detector. For SCC a threshold of 500 × 10³ cells/ml was used.

For both EC and SCC considerable variation was found between herds for sensitivity (the proportion of quarters infected with a major pathogen and detected as abnormal), and specificity (the proportion of quarters free from infection and detected as normal). The mean sensitivity and specificity of EC for the three herds was 49% and 79% respectively, whereas for SCC the means were 71% and 81% respectively.

The variation in sensitivity and specificity of EC between herds was attributed to differences in the distribution of EC for quarters of similar infection status. It was concluded that these differences in herd EC precluded the use of pre-determined EC thresholds which were applicable for detecting mastitis in all herds.     

Loss of reserves of Cu in liver when Cu supplements are withdrawn from dairy herds in the Waikato region

Abstract

AIM: To monitor the consequences of withdrawing mineral Cu supplements from two dairy herds with initially high concentrations of Cu in liver.

METHODS: Two herds were selected from dairy farms in the Waikato region of New Zealand that participated in an earlier survey of Cu supplementation practices and Cu status of dairy cows. The herds were fed pasture, grass and maize silage, plus palm kernel expeller (PKE) containing 25–30 mg Cu/kg dry matter (DM) fed at 2–4 kg/cow/day. No mineral Cu supplements were supplied from January 2009. Pasture samples were collected for mineral analysis in September 2008 and April 2009. Concentration of Cu in liver biopsies from the same 9–10 cows per herd was measured on three occasions between April 2009 and May 2010.

RESULTS: Pastures on both farms contained 10 mg Cu/kg DM, 0.1–0.5 mg Mo/kg DM and 3.5–4.0 g S/kg DM. The initial herd mean concentrations of Cu in liver were 1,500 (SD 590) and 1,250 (SD 640) μmol Cu/kg fresh tissue. In the absence of mineral Cu supplements, those mean concentrations decreased over 12 months to 705 (SD 370) and 1,120 (SD 560) μmol Cu/kg fresh tissue, respectively. For cows in the first herd, the rate of depletion of liver Cu reserves was influenced by initial concentration of Cu, such that high concentration led to faster loss according to first-order kinetics.

CONCLUSIONS: Mineral Cu supplementation was not necessary over 12 months for two dairy herds with mean concentrations of Cu in liver >1,250 μmol Cu/kg fresh tissue, grazing pastures containing 10 mg Cu/kg DM and concentrations of Mo <1 mg/kg DM. The quantity and particularly the duration of feeding PKE appeared to be a factor in whether or not the herd lost substantial reserves of Cu in liver during the year. However, the Cu status of both herds in this study was more than adequate to support late pregnancy and mating.

CLINICAL REVELANCE: Copper status of the herd should be monitored and on-farm management of Cu nutrition should take into account all sources contributing to daily intake of Cu. Where Cu supplementation has been excessive and there is risk of chronic Cu toxicity, mineral Cu supplements may be withdrawn for a period commensurate with the expected rate of liver Cu depletion.     

Impact of molybdenum on the copper status of red deer (Cervus elaphus)

AIM: To determine the effect of increasing molybdenum (Mo) intakes on serum and liver copper (Cu) concentrations and growth rates of grazing red deer (Cervus elaphus). METHODS: Molybdenum- and Cu-amended fertilisers were applied to six 1.1-ha paddocks in a 3 x 2 design. Three levels of Mo were applied on two paddocks at each level in mid April (designated Day 1); levels were: none (control), 0.5 (medium) and 1.0 (high) kg Mo/ha as sodium molybdate. In late May (Day 39), two levels of Cu (none and 3.0 kg Cu/ha, as copper sulphate) were applied to each of the three levels of Mo-treated paddocks. Pasture Mo, Cu and sulphur (S) concentrations were measured at about fortnightly intervals. In late June (Day 74), ten 6-month-old red deer hinds were placed on the six experimental pastures, and serum and liver Cu concentrations were monitored at about monthly intervals for 102 days. The hinds were weighed on four occasions during the trial. RESULTS: Mean pasture Mo concentrations on Day 56 were 2, 4.6 and 11.3 mg/kg dry matter (DM) for the untreated control, medium and high Mo-treated pastures, respectively. Pasture Cu concentration was 95 mg/kg DM on Day 59, 53 mg/kg DM on Day 90, and 9 mg/kg DM by Day 153. Mean S concentration in pasture was 3.3 (range 3.03-3.45) g/kg DM. Copper application to pasture had no significant effect on serum and liver Cu concentrations in deer so data were pooled within Mo treatment. Mean initial (Day 74) serum Cu concentration was 9.2 micromol/L. In the deer grazing the control Mo pasture, this increased to 10.3 micromol/L on Day 112, before decreasing to 6.4 micromol/L on Day 176. In deer grazing the medium and high Mo-treated pastures, mean serum Cu concentrations were 3.8 and 3.9 micromol/L, respectively, on Day 112, and 2.5 and 3.3 micromol/L, respectively, on Day 176. Mean initial (Day 74) liver Cu concentration was 131 micromol/kg fresh tissue. In the deer grazing the control Mo pasture, this declined to 120 and 52 micromol/kg on Days 112 and Day 176, respectively. In deer grazing the medium and high Mo-treated pastures, liver Cu concentrations decreased to 55 and 52 micromol/kg fresh tissue, respectively, on Day 112, and 21 and 20 micromol/kg fresh tissue, respectively, on Day 176. Mean serum and liver Cu concentrations were not significantly different between deer grazing the medium and high Mo-treated pastures, and were lower (serum p=0.003, liver p<0.001) in those groups than in deer grazing the untreated control pastures. No clinical signs of Cu deficiency associated with lameness were observed. Deer grazing pastures that had Mo concentrations >10 mg/kg DM had lower (p=0.002) growth rates (100 vs 130 g/day) than those on pastures containing <2.4 mg Mo/kg DM. CONCLUSION: Increasing pasture Mo concentrations from 2 mg/kg DM to > or =4.6 mg/kg DM significantly reduced serum and liver Cu concentrations in grazing deer. Reduced growth rate was observed at pasture Mo concentrations >10 mg/kg DM.     

Copper, zinc and manganese concentrations in equine liver, kidney and plasma

Five groups of horses were fed different diets of known trace mineral concentration for a minimum of six months. Copper (Cu), zinc (Zn) and manganese (Mn) concentrations were measured in livers of 125 yearling horses and kidneys of 81 yearling horses as an assessment of trace mineral status. Plasma Cu and Zn determinations were made for all horses.

Mean hepatic Cu concentrations of horses fed diets containing 6.9 to 15.2 mg Cu/kg dry matter (DM) feed were 0.27 to 0.33 μmol/g DM tissue. Plasma Cu concentrations ranged between 22.8 to 28.3 μmol/L. There was no simple mathematical relationship between plasma and hepatic Cu concentrations. Mean hepatic Zn concentrations in horses fed diets containing 25.6 to 52.2 mg Zn/kg DM feed were determined to be between 2.75 to 2.91 μmol/g DM tissue. Mean plasma Zn concentrations in groups of horses were between 11.7 to 13.5 μmol/L. Plasma Zn concentrations were not indicative of hepatic Zn concentration. Hepatic Mn concentrations ranged between 0.13 and 0.14 μmol/g DM tissue.

Renal Zn concentrations ranged between 1.55 to 1.63 μmol/g DM tissue and did not differ with diet. Mean renal Mn concentrations were 0.09 μmol/g DM tissue for all groups of horses. Renal Cu concentrations ranged from 0.36 to 0.47 μmol/g DM tissue and differed with diet.

     

Prediction of Serum IgG1 Concentration in Holstein Calves Using Serum Gamma Glutamyltransferase Activity

We examined the relationship between serum gamma glutamyltransferase (GGT) activity and serum gamma immunoglobulin G (IgG1) concentration in Holstein calves. Blood samples were collected from calves aged 1 to 3 days. A follow-up sample was obtained from each calf 2, 7, or 15 days after the initial sampling. Serum GGT activity and lgG1 concentration were measured. Regression models were used to predict lgG1 concentration as a function of age and serum GGT activity measured 2, 7, or 15 days later.
Serum GGT activity and calf age at the time of the second sample were directly related to serum IgG1 concentration in the initial sample in calves aged 3 to 17 days ( r = .54) and in calves aged 3 to 10 days ( r = .63). Models were used to estimate the serum GGT activity equivalent to a serum IgG1 concentration of 1,000 mg/dL. One-day-old calves should have serum GGT activities > 200 IU/L. Four-day-old calves should have serum GGT activities > 100 IU/L. One-week-old calves should have serum GGT activities > 75 IU/L. Calves with serum GGT activities < 50 IU/L should be classified as having failure of passive transfer.     

Non-specific seroreactions against Brucella abortus in ruminants in New Zealand and the presence of Yersinia enterocolitica 0:9

The level of non-specific reactions found in the brucellosis serology of ruminants in New Zealand was very low until July 1992. This changed when, in an export consignment of 1071 deer, 35% reacted in the Brucella abortus tube agglutination test with titres varying from 50 to 200 IU. The reactors were also positive in the Rose-Bengal agglutination test and most of them reacted in the complement fixation test with titres varying from 10 to 80 IU. Yersinia enterocolitica 0:9 was later isolated from one deer of this consignment. It was the first isolate of this serotype recovered in New Zealand from an animal. Shortly after, false reactors occurred more frequently than before in sera from Brucella abortus accredited free cattle herds. As the involvement of Yersinia enterocolitica 0:9 was suspected in these cases, faecal samples from reactors and in-contact animals were cultured for this organism. Yersinia enterocolitica 0:9 was isolated from nine of 19 herds showing one or more false Brucella abortus seroreactions.

Prior to 1990, Yersinia enterocolitica serotype 0:9 had not been isolated in New Zealand, despite the recovery of a number of other bio— or serotypes of the organism from humans and animals. From 1990 onward, serotype 0:9 began to be isolated from human faecal samples with increasing frequency. Since the first isolations from deer and cattle in 1992, it has now also been recovered from a cat and an alpaca and from cattle without any association with false positive Brucella abortus reactions. All serotype 0:9 isolates were of biotype 2.     

An evaluation of the effect of clotting and processing of blood samples on the recovery of copper from bovine blood

The reference ranges for blood copper (Cu) concentration used by SAC and the UK Veterinary Laboratories Agency have been developed for use with plasma only. Copper sequestration into the clot, primarily in the form of caeruloplasmin, means that serum Cu concentrations are significantly lower than plasma Cu concentrations in the same animals. Serum Cu is commonly used as a surrogate for plasma Cu by applying a proportional conversion factor. However the results of these two studies, which involved a total of 104 pairs of serum and Cu samples from 84 Holstein cattle in one herd, suggest that serum Cu concentrations should not be used in this way. First, the reduction in Cu concentration during clotting was unrelated to initial plasma concentration (varying from 8-59%), which suggests that using a proportional conversion factor is incorrect. Secondly, the correlation between plasma and serum Cu concentration, while significant, was relatively low (r2 proportional 0.6), with the individual variation in Cu loss during clotting ranging from 1.5 to 10.8 micromol/L (mean 4.9 micromol/L). Although this study was undertaken using samples from normocupraemic lactating Holstein cattle in one herd, the data strongly suggest that plasma rather than serum Cu should be used to estimate Cu status unless the laboratory used has developed a reliable serum reference range. We also found that factors other than plasma Cu concentration had a significant effect on loss of Cu during the clotting process. However, no significant effect on the loss of Cu during clotting or by varying the incubation/storage time and temperature of bloods prior to clot removal was noted, nor was there any significant effect of age or Cu intake. Furthermore, when the loss of Cu during clotting was examined in the same heifers at two different timepoints, no correlation between the results was found. Further research is required to establish the factors that affect the loss of Cu during clotting.     

Influence of Minerals on the Aetiology of Geophagia in Periurban Dairy Cattle in the Derived Savannah of Nigeria

The aetiology of geophagia in periurban dairy cattle in Nigeria was assessed in relation to the mineral status of the serum, the herbage consumed, and the soil from where the herbage was consumed. The study was carried out using nine herds in the derived savannah zone during both the wet and dry seasons. Generally, the Na, Ca, Fe, Zn and Cu levels in the samples were adequate. However, the forages were low in Ca and the serum was deficient in P. Forage, soil and termite hill samples were also deficient in P. The low levels of P in the soil and termite hill samples showed that the consumption of sand by the cattle did not have any positive impact on their P intake. A seasonal effect was apparent (p<0.05) on the serum Na⁺, PO₄ ^3–, Cu²⁺ and Fe²⁺ concentrations but not on Ca²⁺ and Zn²⁺ concentrations. A seasonal effect was also significant (p<0.05) on the minerals in the forage. There were also seasonal differences in most of the soil and termite hill minerals. Most samples had higher (p<0.05) mineral levels in the dry season. Herd size and farming activity had no apparent influence on the aetiology of geophagia as they did not affect (p>0.05) the concentration of P in the serum, the reported cause of geophagia. Geophagia is probably caused by a marginal level of P in the serum and low levels in the forage and soil. Experimental trials will be required to confirm these observations.     

The effects of levamisole on oxidative stress induced by copper intoxication in broilers

Abstract

AIM: To determine the effects of Cu and levamisole on concentrations of Cu and Fe in plasma and liver, and the effects of levamisole on lipid peroxidation induced by Cu intoxication in broiler chickens.

METHODS: In a 2×4 factorial study, 80 one-day-old Ross PM3 broiler chicks were fed diets for 21 days containing either 8 mg/kg Cu (Low Cu) or 250 mg/kg Cu (High Cu) and were treated with 0 (L0), 4 (L4), 8 (L8) or 16 (L16) mg/kg bodyweight levamisole per day from Day 7 of the study, on three consecutive days in their drinking water. This treatment was repeated three times, at 3-day intervals. On Day 21, blood samples were collected from each bird for analysis of concentrations of Cu, Fe and malondialdehyde, and activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT), superoxide dismutase, catalase and glutathione peroxidase (GSH-Px). The birds were killed and liver samples collected for analysis of Cu and Fe.

RESULTS: Mean concentrations of Cu and Fe in plasma, and Cu in liver, were increased overall in the High Cu groups compared with the Low Cu groups (p<0.001). Compared with the L0 treatment group on the High Cu diet, treatments L4, L8 and L16 decreased concentrations of Cu in plasma, and L8 and L16 increased concentration of Cu in liver (p<0.05). Mean activities of AST and ALT were increased in untreated birds (L0) fed the High compared with Low Cu diets (p<0.01). In birds receiving the High Cu diet, treatments L4 and L8 decreased activities of AST, and L4 and L16 decreased activity of ALT, compared with L0 (p<0.05). The High Cu diet induced an oxidative stress characterised by increased mean concentrations of malondialdehyde and decreased activities of superoxide dismutase, catalase and GSH-Px (p<0.001). Concentration of malondialdehyde, and activities of superoxide dismutase and catalase were not changed following levamisole treatment in birds on the High Cu diet, and activity of GSH-Px was decreased by the L4 and L8 treatments compared with the L0 group.

CONCLUSIONS AND CLINICAL RELEVANCE: The results of the study suggest that treatment with levamisole might alleviate the harmful effects of Cu on the liver, as demonstrated by decreased activities of AST and ALT induced by a diet containing 250 mg/kg Cu.     

补铜对奶牛血浆铜含量和铜蓝蛋白活性及其相关性的影响

本试验选取了大庆地区某奶牛群11头泌乳奶牛，检测补铜前后奶牛血浆铜含量和铜蓝蛋白(CP)活性的变化，同时分析了两者相关性。结果表明：补铜前血浆铜在10.93 μmol/L以下者占被检牛数的54.55%（6/11），其血浆CP活性降低，两者呈现中度相关性（R2=0.53）；补铜后血浆铜含量与CP活性均极显著增高（P<0.01），血浆铜含量达到正常范围，两者呈现低相关性（R2=0.12）。〖JP〗结论：该奶牛场泌乳奶牛群中存在低铜血症病牛，血浆铜蓝蛋白可以作为奶牛低铜血症的一项检测指标。     

Diagnosis of tuberculosis due to Mycobacterium bovis in New Zealand red deer (Cervus elaphus) using a composite blood test and antibody assays

A blood test for tuberculosis in deer was developed as an ancillary test to clarify the status of skin test-positive deer, with non-specific sensitisation following exposure to saprophytic mycobacteria. The blood test incorporates the measurement of the relative humoral and cellular immunological responses to Mycobacterium bovis and M. avium antigens to provia composite test with high levels of sensitivity (>95%) and specificity (>98%). The specificity of the test has allowed it to be used in parallel with the skin test to salvage thousands of tuberculosis-free deer with non-specific skin test-positive reactions, while its high sensitivity has consistently identified M. bovis-specific reactivity in tuberculous skin test-positive animals. The rules for establishing the diagnostic parameters for the cellular and antibody assays were developed by retrospective analysis of the laboratory results using blood samples from many thousand tuberculous or disease-free deer. The sensitivity of the blood test was tested in this study using 150 animals with tuberculosis diagnosed by the isolation of M. bovis. It had sensitivity values of 95.7–95.9% in herds with a low (<2.0% ) or a high (>30.0%) incidence of tuberculosis. The test had a specificity of 98.0% when tested on 218 disease-free animals, 118 of which were skin test-positive.

An antibody test was developed to diagnose M. bovis in skin test-negative “anergic” deer from tuberculosis infected herds. When this test was used with deer blood taken 10 days after reading the skin test, it had a sensitivity of 85.3% for 102 M. bovis-positive deer. When used in combination with skin test, the antibody test complemented the skin test to raise the sensitivity of the combined tests to 95.0%) when antibody-positive or skin test-positive tests were used to diagnose tuberculosis. The specificity of the antibody test was 100% when used to evaluate 218 disease-free deer from non-infected herds.     

Studies on the incidence of clinical mastitis and blood levels of vitamin E and selenium in dairy herds in England

In a trial conducted in the south of England in January to February 1989, blood samples were obtained from nine dairy herds with more than 30 cases of clinical mastitis/100 cows and from nine herds with less than 30 cases/100 cows during the previous 12 months. Whole blood glutathione peroxidase (GSHPx) activity and plasma vitamin E concentration were determined for 12 cows in each herd. The mean (+/-sd) values for the herds with the lower incidence of mastitis were 7.57 +/- 1.86 micrograms/ml plasma vitamin E and 23.8 +/- 22.8 U/ml rbc GSHPx activity, compared with 7.74 +/- 1.69 micrograms/ml plasma vitamin E and 20.61 +/- 8.8 U/ml rbc GSHPx activity for the herds with the higher incidence of the disease. These values indicate that the vitamin E levels were generally adequate but that some animals and herds had low GSHPx activities, suggesting that their diets may have contained inadequate selenium. The activities of GSHPx and the vitamin E levels in plasma were not significantly different in the two groups of herds, and no relationship was found between the two nutrients and the incidence of clinical mastitis. However, there was a significant negative correlation between the activity of GSHPx and the bulk milk cell counts in the herds with a low incidence of mastitis suggesting that there was an association between the incidence of subclinical mastitis or inflammation and the selenium status of these herds.