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Small Ruminant Research 68 (2007) 279–284

Macromineral status of grazing sheep in asemi-arid region of Pakistan

Z.I. Khan a, A. Hussain a, M. Ashraf a,M.Y. Ashraf b, L.R. McDowell c,∗

a Department of Botany, University of Agriculture, Faisalabad, Pakistanb Nuclear Institute for Agriculture and Biology (NIAB) Faisalabad, Pakistan

c Animal Sciences Department, University of Florida, Building 459, Shealy Drive,P.O. Box 110910, Gainesville, FL 32611-0910, USA

Received 14 February 2005; received in revised form 1 November 2005; accepted 1 November 2005Available online 27 December 2005

bstract

The present study was conducted to evaluate seasonal and animal class effects on the macromineral status of lactating andon-lactating (female) classes of grazing Thalli sheep in a semi-arid region of the south western Punjab, Pakistan, on the basis ofineral concentrations in blood plasma. There were 20 animals in each class. Plasma Ca, Na, and K in both classes of sheep showed

ffects of season (P < 0.05), animal class (P < 0.001), and interaction by season and animal class (P < 0.01) whereas, plasma Mghowed only interaction effect between season and animal class (P < 0.05). Moderately deficient levels (60–80% plasma samplesere below the normal range) were found for sheep at this age and weight for Ca during winter and of K and Na during summer.

n contrast, levels of Ca in summer, that of Na and K during winter, and of Mg during both seasons, were marginally deficient80–100% plasma samples were below the normal range) in lactating sheep. While in non-lactating sheep moderately deficient

evels of Ca in winter, Na in summer, and marginally deficient levels of K and Mg during both seasons were found on the basis ofritical levels of these macrominerals in plasma. Overall macromineral status of these sheep based on plasma concentrations maye considered deficient. From these blood analyses, it was concluded that macromineral status of sheep on this specific ranch inunjab needs supplementation of macrominerals with the implication of similar needs for other regions of Pakistan.2005 Elsevier B.V. All rights reserved.

eywords: Macromineral; Grazing sheep; Blood; Status; Pakistan

. Introduction

The assessment of mineral needs of animals hasome to include determination of the minerals in theissues, fluids, and products as well as such gross crite-

∗ Corresponding author. Tel.: +1 352 392 7561;ax: +1 352 392 7652.

E-mail addresses: drzafar10@hotmail.com (Z.I. Khan), mcdow-ll@animal.ufl.edu (L.R. McDowell).

921-4488/$ – see front matter © 2005 Elsevier B.V. All rights reserved.doi:10.1016/j.smallrumres.2005.11.003

ria as weight gains, milk yield, etc. Many factors affectrequirements, including kind and levels of production,age, level, and chemical form of elements, interrelation-ships with other nutrients, mineral intake, breed, andanimal adaptation. The detection of mineral deficienciesor excesses involves clinical, pathological, and analyt-ical criteria as well as response from specific element

supplementation. Clinical signs of mineral deficienciesalong with soil, water, plant, and animal tissue analy-ses have all been used with varying degrees of successto establish mineral deficiencies and toxicities (Mills,

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1987; Underwood and Suttle, 1999; McDowell, 1997,2003).

For several decades, a major goal in mineral researchhas been to discover and/or develop simple and accuratebiochemical measurements of the status of animals forthe minerals in which there are important practical prob-lems (Miller and Stake, 1974). Like soil and plants, ani-mal tissue or fluid mineral concentrations are influencedby many factors. Nevertheless, when appropriate inter-pretation is made, animal tissue or fluid concentrationsare often better indicators of the mineral status of live-stock than either plant or soil concentrations (McDowell,1997, 2003).

In many world regions the grazing ruminants oftendo not receive mineral supplements except for commonsalt, and a free-choice complete mineral mixture is rec-ommended as an insurance for providing minerals wheredietary concentrations are unknown or highly variabledue to season, location, forage species and animal poten-tial (McDowell, 2003). It is important to determine themineral profile based on animal fluid/tissue concentra-tions in order to estimate the mineral needs of grazingruminants, as well as the time of the year when they aremost needed.

Unfortunately, no tissue or fluid will portray thestatus of all minerals. Blood, urine, saliva, and milkmay be easily sampled with a minimum of time andwithout sacrificing the experimental animal. Amongother fluids, blood plasma is an indicator of the sta-tus of certain elements and levels of these elementsin plasma are of value in the determination of defi-ciency or toxicity states. However, individual vari-ability can be very high and external contaminationmay provide problems for certain trace element statusevaluation.

The objective of this study was to evaluate and com-pare macromineral status of two classes of sheep grazingsimilar pastures on the basis of mineral concentrationsin blood plasma.

2. Materials and methods

2.1. Study area

The study was conducted at the Pakistan LivestockExperimental Station using a herd of Thalli lactating andnon-lactating sheep (female), grazing similar pastures onthe farm during summer and winter of 2003, correspond-

ing to the wet and dry seasons. The farm “LivestockExperimental Station Rakh Khaire Wala” is located inSouthern Punjab, Pakistan. Pastures were predominantlygrasses, legumes, tree leaves and crop wastes. In addi-

esearch 68 (2007) 279–284

tion, all animals on the farm had access at all times to afree-choice complete mineral mixture. This sheep ranchcomprises 400 ha and receives annual precipitation of250–750 mm, mainly restricted to July and August. Thesoils are sandy and vertisols. The ranch has about 7000animals of which 2000 are breeding sheep. This ranch isin a low-lying semi-arid region of south western Punjabbetween latitudes 23◦ and 36◦N and longitudes 60–75◦E.Average temperature during the experimental year wasbetween 38 ± 5 ◦C during summer and 15 ± 7 ◦C duringwinter, relative humidity 48 ± 5% during summer and80 ± 8% during winter.

The livestock farm was characterized by two pastures,denoted as feeding sites, one pasture was intensivelymanaged with fertilized soils, irrigated with canalwater, and with grazing reserves characterized by theavailability of sown forage species including Panicum,Andropogon, Penisetum, Setaria, Medicago sativa,Trifolium alexandrium, Hordeum vulgare, Cichoriumintybus, Cynodon genera, Vernal grass, Imported velvetgrass, Tall fescue, Orchard grass, Molasses grass,Elephant grass, Pangola grass, and Jaragua grass. Theother pasture with unfertilized soils, was a barren anduncultivated area with natural weed-like vegetation andlow intensity cropping largely accessible to grazinganimals. This pasture was overgrazed with extensivereplacement of perennial grasses by annual grasses,and bush encroachment by Accacia spp., Ziziphusmucronata, Trachipogon spp., Cyperus rotandus,Tribulus terrestris, Chenopodium morale, Lathyrusodoratus, Alhagi spp., Salavadora spp., and Calotropisspp. Each pasture maintained 3 animals/ha/year under arotational grazing system.

2.2. Animals

Twenty each of clinically healthy lactating and non-lactating sheep were used for the study purpose. Averagebody weights were 40–45 kg. These animals were pre-dominantly of this Thalli breed with variable degrees ofcross breeding. The lactating sheep were in their sec-ond lactation. All experimental animals were the samethroughout the study period.

2.3. Housing and management

The work was carried out on the farm for blood collec-tion. The lactating and non-lactating sheep were housed

in a well-ventilated building which was divided by solidwood walls into pens, provided with straw bedding,30 cm thickness, and supplied with fresh clean watercontaining buckets, feed troughs, and salt blocks, for the

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ree access to the lick. All hygienic measures were goodor the farm.

.4. Feeding and nutrition of animals

This work was carried out under two different feed-ng systems which are stable-diet (indoors) and grazingoutdoors) conditions according to the season of the year.he animals were fed the stable diets formulated to cover

heir nutrient requirements (NRC, 1985). However, bothnimal classes were gradually transferred to the grassasture for grazing. The time of grazing was 2.5–5.0 h.he animals were on the grass pasture ad libitum andupplemented with a free-choice mineral mixture. Sam-les of feed ingredients, and grass pasture were collectederiodically, dried, and analyzed for the proximate chem-cal composition (Khan, 2003) with overall means ofummer and winter seasons, respectively, as follows: Ca0.37; 0.30%), Mg (0.20; 0.17%), Na (0.05; 0.03%), K3.1; 0.6%), Cu (4.6; 5.8 ppm), Fe (60; 85 ppm), Mn (68;5 ppm), Zn (25; 30 ppm), Co (0.125; 0.145 ppm), ande (0.045; 0.089 ppm). The free-choice mineral mixturead the following composition: Ca 15.7%, P 6.3%, Mg.0%, K 0.3%, Na, 11%, Mn 2400 ppm, Fe 6000 ppm,n 2630 ppm, Cu 20 ppm, Co 16.4 ppm, Mo 10.8 ppm,nd Se 0.8 ppm.

.5. Sample collection and preparation

About 10 ml blood samples were collected four timesuring each season from 20-lactating, and 20-non-

able 1acromineral concentrations in plasma of lactating and non-lactating sheep a

ariable mg/L and significance ofeason or animal class

Significance of season oranimal class

Sea

a2+, C.L.:80 S*, A***, SA** WinSum

g2+, C.L.:20 S:ns, A:ns, SA* WinSum

a+, C.L.:3000 S**, A***, SA* WinSum

+, C.L.:200 S*, A***, SA* WinSum

eans are based on 80 samples in each season for each class of animals. S: se.L. (critical concentrations) values: McDowell et al. (1984) (mg/L), Ca (80)* Significant at 0.5 level.

** Significant at 0.01 level.** Significant at 0.001 level.

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lactating sheep by jugular vein puncture with a syringeand needle, then transferred into evacuated tubes con-taining lithium heparin as an anticoagulant. For analyt-ical preparation, blood samples were centrifuged at aspeed of 300 rpm for 20 min and plasma was harvestedinto polyethylene tubes. The supernatant plasma wasdecanted into crucibles and was first dried in a forceddraught oven at 60 ◦C for 6 h. It was then ashed in amuffle furnace at 550 ◦C followed by acid digestion toprepare ash solutions (Fick et al., 1979; Mpofu et al.,1999). The ash solutions were then analyzed for Ca, Mg,Na, and K using an atomic absorption spectrophotometer(Perkin-Elmer, 1984).

2.6. Statistical analysis

The data thus obtained during the study were analyzedby Statistical Analysis System (SAS for Windows V9;SAS Inst., Inc., Cary, NC). Animal blood plasma sam-ples were analyzed as split-plot design (Steel and Torrie,1980), with animal class as the main plot and season assub plot. Significance levels ranged from 0.05 to 0.001.Differences among means were ranked using Duncan’sNew Multiple Range Test (Duncan, 1955).

3. Results and discussion

Both lactating and non-lactating sheep plasma Caconcentrations were affected by seasons (P < 0.05), ani-mal class (P < 0.001) and interaction of season by animalclass (P < 0.01) (Table 1).

s related to season and animal class

son Animal class

Lactating sheep Non-lactating sheep

Mean S.E. Mean S.E.

ter 60.5 1.56 63.6 1.29mer 76.7 1.40 91.78 0.61

ter 19.4 0.43 19.2 0.33mer 21.36 0.26 21.54 0.34

ter 2589 38.30 2759 22.7mer 2383 31.0 2277 21.7

ter 177 3.31 171 2.7mer 158 2.9 195 3.3

ason; A: animal; SA: season × animal interaction. ns: non-significant., and Mg (20); Miles et al. (2001) (mg/L), Na (3000), and K (200).

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Plasma Ca from both classes of sheep were higherin summer, but non-lactating sheep plasma containedhigher (P < 0.001) Ca during both seasons than that oflactating sheep. The plasma Ca concentrations in lactat-ing sheep were numerically lower than in non-lactatingsheep on an overall basis. Similarly, plasma Ca of lactat-ing animals were lower during winter than in summer.But in non-lactating sheep, plasma Ca was higher dur-ing summer than during winter. Overall, the plasma Cain non-lactating sheep was higher during both seasonsthan that in lactating animals. However, these values inboth groups of animals, except in non-lactating sheepduring summer, were lower than the standard Ca refer-ence value of (80 mg/L) (McDowell et al., 1984). Thesefindings were in accordance with those reported earlier(Lee et al., 1978; Kulkarni and Talvelkar, 1984). In lac-tating sheep, the Ca homeostasis is maintained by anincreased rate of dietary Ca absorption, decreased rateof urinary Ca excretion, and mobilization of bone Caunder the complex physiological action of parathyroidhormone, calcitonin, and vitamin D (McDowell, 2003).

Plasma Ca in lactating sheep during summer and innon-lactating animals during winter fell in-between val-ues reported by different researchers in earlier studies(Seshiah, 1962; Husnain et al., 1981; Nemat et al., 1983;Kulkarni and Talvelkar, 1984), but lower than from otherruminant research groups (Lakke, 1954; Areadio et al.,1956; Sheikh, 1975; Shukla et al., 1980; Guay, 1988).

There were no obvious season or animal class(P > 0.05) effects for plasma Mg concentrations for bothlactating and non-lactating sheep, but interaction ofseason and animal class was found to be significant(P < 0.05) (Table 1). During winter, plasma Mg concen-trations were non-statistically lower in both classes ofsheep than during summer.

All plasma Mg samples for both animal classes dur-ing winter were deficient in Mg but none in summer.Plasma Mg in sheep were lower than the reference valueof 20 mg/L presented by McDowell et al. (1984) onlyduring winter, plasma Mg concentrations for both sheepclasses were higher than the suggested critical valueduring summer. Mean plasma Mg in lactating and non-lactating sheep was similar. Based on the suggested crit-ical value of 20 mg/L (McDowell et al., 1984), plasmaMg was marginally deficient in both classes of sheep.The differences in plasma Mg levels of lactating sheepand non-lactating sheep on the basis of seasonal changeswere non-significant. The highest value of plasma Mg

was found in both lactating and non-lactating sheepduring summer, but plasma Mg in all samples of lac-tating sheep during summer and in non-lactating sheepduring winter fell in-between these values. Plasma Mg

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concentrations in this study are not in line with someinvestigations (Sharma and Prasad, 1982; Limpoka etal., 1987; Oba et al., 1988), who reported plasma Mgconcentrations much higher than the present findings.This may be attributed to different management, nutri-tion, animal type, and various other factors involved inthe mineral absorption from the gastro-intestinal tractof animals (McDowell, 1985). However, the findings ofthe present study are in agreement with other research(Kiatoko et al., 1978; McDowell et al., 1987; Tejada etal., 1987; Iqbal et al., 1988).

In lactating and non-lactating sheep plasma Na con-centrations were affected by both season (P < 0.01) andanimal class (P < 0.001) and the interaction of seasonclass (P < 0.05) (Table 1). Generally, plasma Na concen-trations in both groups of animals were higher duringwinter than summer. Plasma Na was below the referencevalue of 3000 mg/L (Miles et al., 2001) in all samples.

Mean plasma Na of lactating sheep was lower thanthat in non-lactating sheep during winter, while thereverse was true for summer. However, these valueswere below the mean plasma Na level suggested byUnderwood (1981), and Miles et al. (2001). Lowerplasma Na in lactating animals is likely due to the highsecretion of Na through milk (Underwood, 1981). Theother minerals of Ca, K, and Mg were not affected asmuch as Na during dietary stress or other environmen-tal fluctuations (wet and dry conditions). These findingswere in agreement with those of Limpoka et al. (1987),who reported similar results while working with ani-mal, grass, and soil mineral concentrations in variousprovinces of Thailand. Plasma Na values recorded inthis study were lower than those of Gillani (1984) andKulkarni and Talvelkar (1984), who reported a highrange of plasma Na concentrations in various dairy ani-mals. This might have been due to different environmen-tal conditions and/or length/stage of lactation, animaltypes or feed resources, which were not specified in thosestudies.

Animal class and seasons for plasma K were sig-nificant (P < 0.05, <0.01), respectively, with no seasonand animal class interaction (P > 0.05) for plasma K(Table 1). Mean plasma K concentrations in lactatingsheep were higher during winter and in non-lactatingsheep during summer. During both seasons in both ani-mal classes plasma K concentrations were below thecritical value of 200 mg/L (Miles et al., 2001). However,when plasma K in both groups of animals was compared,

it was higher in lactating sheep during winter and lowerduring summer than in non-lactating sheep. This vari-ation could have been due to nutritional, managementor stage of lactation and other factors involved in the

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bsorption interactions, antagonisms, and excretion ofinerals in the intestinal tract of animals (Panday andoy, 1968; Kulkarani and Talvelkar, 1984). Plasma Koncentrations in this study are below the normal lev-ls (200 mg/L) and not in agreement with Iqbal et al.1988), Oba et al. (1988), and Ravindra et al. (1988),ho worked with dairy animals. However, these find-

ngs are in agreement with those of Barua et al. (1988),ho reported nearly similar concentrations of plasma K

or various domestic animal classes.The present investigations show that marked differ-

nces in the concentration of all plasma macrominer-ls examined exist between lactating and non-lactatingheep when retained on similar pastures. Variation inlasma macromineral status in different animal classesay be attributed to many factors affecting mineral

equirements which include nature and level of produc-ion, age levels and chemical forms of elements, inter-elationships with other nutrients, mineral intake, classf animals, and animal adaptation (McDowell, 1985).ineral requirements and status are highly dependent

n the level of productivity and physiological state ofhe animal (NRC, 1985). There is extensive evidencehowing marked variations within breeds and betweennimal class, in the efficiency of absorption of min-rals from the diet, 3–35% for Mg in dairy animals,nd 2–10% for Cu in adult sheep (Field, 1984). It haseen observed that when different breeds and classes ofnimals were grazing certain pastures, one breed exhib-ted signs of Cu poisoning, whereas the other showedigns of Cu deficiency. The most possible causes forhis apparent variation in dietary requirements for some

acrominerals between breeds/classes could be geneticifferences in the efficiency of absorption (Field, 1984).nimals most susceptible to some macroelement defi-

iencies are young rapidly growing animals and thoseuring lactation. Dietary requirements may decline withge, because the major requirement for growth oftenemains constant, while appetite increases in proportiono body size.

Specific mineral requirements are difficult to pin pointince exact needs depend on chemical form and numer-us mineral interrelationships. With some elements thehemical form has a major impact on the availabilityf the element (Miller, 1979; Towers and Grace, 1983;race and Lee, 1990).Age and class of animals can affect requirements of

inerals through changes in efficiency of absorption. In

eneral, young animals absorb minerals more efficientlyhan older animals (McDowell, 2003). Grazing habitsnd preferences for different grasses by lactating andon-lactating sheep may be a cause of different plasma

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mineral concentrations found in the present investiga-tions (San Martin and Bryant, 1989).

4. Conclusions

Results from the investigation on macromineral sta-tus of grazing sheep in the specified region of Pakistansuggest that these macrominerals particularly Ca, dur-ing both seasons for lactating and during summer fornon-lactating sheep, Mg during winter for both classes,Na for both classes during winter and summer and Kduring winter for both classes and during summer onlyfor non-lactating sheep were definitely deficient. There-fore, supplementation of these ruminant groups with abioavailable mineral mixture is needed to enhance theblood level of these essential minerals. However, furtherresearch is needed to determine if mineral deficienciesare limiting grazing livestock production.

Acknowledgement

This research is Florida Agric. Exp. Sta. No. R-10644.

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