Interntionl Conference: Septemer 15-17, 2008 Rgus - Itly Innovtion Technology to Empower Sfety, Helth nd Welfre in Agriculture nd Agro-food Systems Strtegies to reduce het stress in sheep housing Croprese M. 1, Alenzio M., Mrino R., Muscio A., Sntillo A., Sevi A. 1 Diprtimento PRIME nd Istituto per l Ricerc e le Appliczioni Biotecnologiche per l Sicurezz e l Vlorizzzione dei Prodotti Tipici e di Qulità (BIOAGROMED), Università di Foggi, vi Npoli, 25, 71100 Foggi, Itly. Tel ++39-0881-589332, Fx ++39-0881- 589331, e-mil m.croprese@unifg.it Astrct Three experiments were crried out to ssess the effects of ventiltion regimens, shding nd time of feeding on the welfre nd production performnce of lctting ewes in summer in Mediterrnen climte. In the first tril we tested the effects of three different ventiltion regimens: low (33 m 3 /h/hed; LVR), moderte (66 m 3 /h/hed; MVR) nd progrmmed (operting t 30 C ir temperture nd 70% reltive humidity; PROGR). In the sheep houses fter LVR nd MVR tretments were detected higher ir concentrtions of mmoni nd cron dioxide thn fter PROGR tretment nd sheep of those groups showed higher respirtion rte nd rectl tempertures. Ewes under the LVR tretment lso displyed impired humorl immune responses nd higher plsm cortisol levels thn ewes under PROGR or MVR tretment. In the second tril, we tested the effects of two ventiltion rtes (35 vs 70 m 3 /h per ewe), two ir-speeds (2 vs 4 m/s) nd two lengths of ventiltion cycles (30 vs 60 min/cycle). Results suggest tht proper ir speed nd length of ventiltion cycles, side from ventiltion rte, re required to sustin welfre of housed sheep in summer. When compring the effects of providing shded res we found tht sheep exposed to solr rdition displyed enhnced respirtion rte nd incresed rectl temperture compred with nimls protected from solr rdition. Exposure to solr rdition lso depressed cellulr immune response nd dversely ffected the hygienic qulity of ewe milk In ddition, sheep exposed to direct solr rdition exhiited n increse of inctive ehviours nd showed high non-esterified ftty cids plsm concentrtions. Keywords : sheep welfre; het stress; solr rdition; ventiltion regimen. Introduction Wide res of the Mediterrnen sin, where diry sheep re very common, hve mient tempertures in lte spring nd summer tht often exceed the therml neutrl zone (5 to 25 C) of sheep (Curtis, 1983). Hot climtes my induce rise in ody temperture nd reth rte, which increses energy requirements for mintennce y 7-25%. Feed intke decreses in het stressed sheep (Adll et l., 1993), especilly when they re offered low qulity feed (Cost et l., 1992), due to oth the effort of reducing het production nd the slower feed trnsit through the digestive trct. Under these conditions, ody reserves of ft nd nitrogen re used to supply energy through gluconeogenesis t the expense of the mmmry glnd, especilly in erly lctting nimls (Amrl-Phillips et l., 1993). High mient tempertures cn lso result in plsm minerl imlnce, especilly due to reduction in sodium, potssium, clcium nd phosphorus nd increse in chloride concentrtions (Kume et l., 1987; Schneider et l., 1988). Pulished evidence does not prove tht mient temperture is directly relted to udder helth, ut indictes tht it my interct with other predisposing conditions to exert n influence (Klstrup et l., 1987). Beede nd Collier (1986) indicted protection from solr rdition nd improvement of nutritionl mngement s well s use nd genetic development of het-resistnt reeds s the min strtegies to improve
Interntionl Conference: Septemer 15-17, 2008 Rgus - Itly Innovtion Technology to Empower Sfety, Helth nd Welfre in Agriculture nd Agro-food Systems productivity in nimls rised in hot climtes. Ventiltion is one of the most importnt fctors in ffecting the welfre nd production performnce of frmed nimls, too. Indequte ventiltion systems cnnot provide n efficient control of temperture nd humidity in niml houses nd cn led to incresed irorne prticulte nd gseous pollutnt concentrtions, which cn represent significnt urden to the respirtory trct of humns nd livestock (Rylnder, 1986; Hrtung, 1994). Previous experiments hve shown tht poor ventiltion is responsile for incresed eril concentrtions of vile microes, mmoni nd cron dioxide, reduced feed efficiency nd enhnced ggressive interctions in frmed nimls. Three experiments were crried out to ssess the effects of ventiltion regimens, shding nd chnging time of feeding to lte fternoon on the welfre nd production performnce of lctting ewes in summer in Mediterrnen climte. Mteril nd methods The experiments were conducted t Segezi reserch sttion of the Itlin Istituto Sperimentle per l Zootecni (ltitude: 41 27 6 nd longitude: 15 33 5 ). The climte of this re is Mediterrnen, with rinfll of out 500 mm, men temperture of 24.1 C nd men reltive humidity of 59.4% during summer over the lst 40 yers. In the first tril 36 lctting Comisn ewes were divided into three groups of 12 nd seprtely housed on strw litter in 8m x 3m nd 3.5m high rooms of the sme uilding. Ech room ws provided with mechnicl negtive-pressure system of ventiltion, in which 0.28 m 2 suction fns were plced t 2.5 m from the floor nd two 0.36 m 2 ir inlets were plced t the ground level on the opposite wll. Fn speed ws kept constnt t 4.6 m/s. The three groups were designed low (LVR), moderte (MVR) nd progrmmed ventiltion regimen (PROGR). In LVR nd MVR rooms, fns provided 10 ventiltion cycles of 12.5 nd 25 min/h, respectively. In the PROGR room, the fn ws connected to temperture nd humidity sensors, which provided n on/off two stge control function switching power to the fn. Ventiltion system ws progrmmed to operte t 30 C mient temperture nd 70% reltive humidity. A men ventiltion rte of 33, 66 nd 173 m 3 /h per ewe ws provided in LVR, MVR nd PROGR rooms. The mient temperture nd the reltive humidity inside ech room were continuously monitored throughout the tril y mens of thermo-hygrogrphs (LSI) plced t height of 1.5 m from the floor. Air ws smpled 0.6 m from the floor twice week t 0900 (fns switched off in LVR nd MVR rooms) nd t 16.30 h (fns switched on in ll rooms). The concentrtion of mesophilic micro-orgnims, coliforms nd yests/moulds were smpled using Surfce Air System pump (PBI Interntionl, Miln, Itly). Air concentrtions of totl ( > 5 µm) nd rethle ( = 2-5 µm) dust were recorded twice weekly, using Digit pumps (Zmelli, Breggio-Miln, Itly). A Lippmn cyclone ws used for collecting rethle dust. Cellulose nitrte filters hving dimeter of 37 mm nd pore size of 0.8 µm were used for this purpose. Air concentrtions of gseous pollutnts were lso recorded twice week using Gs Detection Pump (Dräger, Sicherheitstechnik Gmh, Lüeck, Germny), provided with cron dioxide, hydrogen sulphide, mmoni nd methne detection tues. Respirtion rte (RR) nd rectl temperture (RT) were monitored in ll nimls throughout the tril. At 1430 RR ws recorded y trined oserver y counting the rte of flnk movement nd soon fter RT ws mesured with n electronic thermometer hving n ccurcy to 0.1 C. Behviorl oservtions were recorded y two trined oservers equipped with video cmers every 15 min from 0930 to 12.30 h once per week. The mesurement criterion t ech oservtion period ws the numer of nimls engged in ech of two postures (stnding or lying) nd of seven ehviorl ctegories, which were eting, drinking, ruminting, wlking, self-grooming nd idling. The phytohemgglutinin (PHA) skin test ws
Interntionl Conference: Septemer 15-17, 2008 Rgus - Itly Innovtion Technology to Empower Sfety, Helth nd Welfre in Agriculture nd Agro-food Systems performed to induce non-specific delyed-type hypersensitivity. At d 3, 20 nd 40 of the experiment, 1 mg of PHA (Sigm-Aldrich Itli, Miln, Itly) dissolved in 1 ml of sterile sline solution ws injected intr-dermlly into the middle of two 2 cm wide circles stmped on shved skin in the upper side of ech shoulder. The skinfold thickness ws determined efore PHA injection nd 24 h fter with cliper. At d 2 of the study, 6 mg of chicken egg lumin (Sigm-Aldrich Itli) dissolved in 1 ml of sterile sline solution nd in 1 ml of incomplete Freund s djuvnt (Sigm Aldrich-Itli) were injected sucutneously in oth shoulders of ech ewes. A second injection without djuvnt ws repeted 9 dys lter. Antiody titer ws determined in lood smples collected in heprinized vcuum tues immeditely efore the first ntigen injection (2 dys) nd then t 11, 21, 30 nd 40 dys of the study period. An ELISA ws performed in 96-well U-ottomed microtiter pltes. At d 37 ewes were intrvenously injected with 2 IU porcine ACTH/kg ody weight 0.75 (Sigm-Aldrich Itli). Blood smples (10 ml) for evlution of cortisol concentrtions were collected in vcuum tues from the jugulr vein immeditely efore nd 1, 2 nd 4 h fter ACTH injection. Hormone concentrtion ws determined y rdioimmunossy specific for ovine cortisol. Ewes were milked twice dily (08.00 nd 15.00 h) using pipeline milking mchines. Milk yield ws recorded dily y mens of grduted mesuring cylinders ttched to individul milking units. Milk smples, consisting of proportionl volumes of morning nd evening milk, were individully collected weekly in 200 ml sterile plstic continers fter clening nd disinfection of tets (70% ethyl lcohol) nd dischrging the first strems of foremilk. Milk smples were crried in our lortory y mens of trnsport tnkers t 4 C. The following mesurements were crried out in greement to Interntionl Diry Federtion stndrds, unless otherwise indicted: ph, totl protein, ft nd lctose content using n i.r. spectrophotometer (Milko Scn 133B; Foss Electric, Hillerød, Denmrk), csein content, somtic cell count (SCC) using Foss Electric Fossomtic 90 cell counter nd polymorphonucler neutrophil leukocyte count (PMNLC), y mens of direct microscopic count in milk smers stined with My-Grünwld-Giems, renneting chrcteristics (clotting time, rte of clot formtion nd clot firmness fter 30 min) using Foss Electric Formgrph. The milk cogulting index (CoI) ws clculted s the clot firmness to clotting time + rte of clot formtion rtio. At the eginning of the tril, nd fortnightly during the study period, the following cteriologicl nlyses were crried out on milk, ccording to Interntionl Diry Federtion stndrds: enumertion of mesophilic cteri, psychrotrophs on plte count gr, totl nd fecl coliforms. The ody weights nd ody condition scores of the ewes (in sixpoint scle with 0=thin nd 5=ft) were recorded during the tril. In the second tril 36 lctting Comisn ewes were divided into three groups of 12 nd seprtely housed in three rooms of the sme uilding s in the first tril. In ll rooms, fns provided 10 ventiltion cycles per dy. Tretments were: low ventiltion (LOV-30), moderte ventiltion with short ventiltion cycles t high ir speed (MOV-30) nd moderte ventiltion with long ventiltion cycles t low ir speed (MOV-60). In the LOV-30 room 30 min ventiltion cycles were provided t fn speed of 2 m/s. In the MOV-30 room 30 min ventiltion cycles were provided t fn speed of 4 m/s, while in the MOV-60 room 60 min ventiltion cycles were provided t fn speed of 2 m/s. In ll rooms, ventiltion rte ws checked dily y plcing hot wire nemometer (LSI, I-20090, Settl Premenugo, Miln, Itly) over the ir outlet nd converting redings to m 3 /h per ewe. The fns provided men ventiltion rte of 35 m 3 /h per ewe in the LOV-30 room nd 70 m 3 /h per ewe in the MOV-30 nd MOV-60 rooms. Mesures tken on ir, nimls nd milk were the sme of the first tril. In the third tril, 40 lte-lctting Comisn ewes were divided into four groups of 10 ech, which were seprtely kept in open 5m x 12m pens with mesh-fence oundries. Shde
Interntionl Conference: Septemer 15-17, 2008 Rgus - Itly Innovtion Technology to Empower Sfety, Helth nd Welfre in Agriculture nd Agro-food Systems ws provided y 3m x 8m nd 3.5 m high rickwork rooms djcent to open pens. During the study, nimls were either exposed (EXP; not offered shde) or protected from solr rdition (PRO; offered shde). For ech solr rdition tretment, feed ws offered either in the morning t 10.00 (EXPM nd PROM groups) or in the fternoon t 16.00 (EXPA nd PROA groups), nd the nimls hd free ccess to the feed t ll times therefter. Refusls were collected nd weighed dily t 10.00, 13.00, 16.00 nd 19.00 h. Open pens in EXP groups nd shded rooms in PRO groups were provided with 2 mngers. Averges of dily DM intke were 2.41, 2.25, 2.39 nd 2.28 kg/ewe in PROM, EXPM, PROA nd EXPA groups, respectively. Wter ws ville t ny time for ll groups from utomtic drinking troughs. Amient temperture nd reltive humidity in protected nd exposed res were monitored throughout the tril. Respirtion rte (RR) nd rectl temperture (RT) were mesured in ll nimls twice weekly. The phytohemgglutinin (PHA) skin test ws performed to induce nonspecific delyed-type hypersensitivity t d 10, 20 nd 32 of the experiment, s descried in previous trils. Behviorl oservtions were recorded y trined oservers equipped with video cmers every 15 min from 08.00 to 20.00 h once per week. Jugulr lood smples were tken from ll ewes t the eginning nd t d 21 nd 42 of the experiment. Blood smpling ws crried out t 07.00 h. Smples were centrifuged for 20 min t 3,500 r.p.m. nd plsm ws stored t -20 C, except for su-smple which ws nlyzed enzymticlly for plsm concentrtions of glucose immeditely. The following metolites nd enzymes in plsm were determined within three dys from lood smpling: totl protein, lumin, iliruin, NEFA, clcium, inorgnic phosphorus, chloride nd mgnesium, using colorimetricl methods, nd totl cholesterol, ure N, cretinin, sprtte mino-trnsferse (AST/GOT), lnine mino-trnsferse (ALT/GPT), gmm-glutmiltrnsferse (γ-gt), lctte- dehydrogense (LDH) nd lkline phosphtse (ALP), using enzymticl methods. Sodium nd potssium were diluted in lithium solution nd mesured y TRIFLAMM flme photometer (Digiflme, 00100 Rome, Itly). Ewes were milked twice dily (08.00 nd 15.00 h) using pipeline milking mchine. Dily milk yield ws recorded nd milk smples were nlyzed s descried in previous trils. The ody weights nd ody condition of the ewes were recorded t the eginning nd t d 21 nd 42 of the study period, fter the morning milking ut efore feeding. All dt were sujected to n nlysis of vrince, using the GLM procedure for repeted mesures in the SAS sttisticl softwre. When significnt effects (P < 0.05) were found the Student s t test ws used to locte significnt differences etween mens. Results First tril. Significntly higher temperture nd humidity (P < 0.001) were found in LVR thn in PROGR room, while oth LVR nd MVR tretment resulted in higher NH 3 nd CO 2 ir concentrtions thn PROGR tretment (P < 0.05). LVR nd MVR ewes hd higher RT thn PROGR ewes (P = 0.001). LVR nimls lso exhiited higher idling compred to PROGR (P < 0.01) nd lower feeding times thn MVR (P < 0.05) nd PROGR nimls (P < 0.01). LVR ewes displyed significnt lower ntiody titers (P = 0.07) thn PROGR nimls (Figure 1) nd higher plsm cortisol levels thn PROGR (P < 0.01) nd MVR ewes (P < 0.05) 60 min fter ACTH injection (Figure 2). LVR tretment resulted in lower yields of milk (P < 0.01) nd reduced feed efficiency (P < 0.01) thn PROGR tretment. Second tril. Averges of mximum THI were out 3 points higher in the LOV-30 nd the MOV-30 thn in the MOV-60 room, wheres no differences emerged in the ir concentrtions of dust, gseous pollutnts nd microorgnisms. Significnt interctions of tretment x time (P < 0.05) were found for respirtion rte, nd for the time the ewes spent
Interntionl Conference: Septemer 15-17, 2008 Rgus - Itly Innovtion Technology to Empower Sfety, Helth nd Welfre in Agriculture nd Agro-food Systems lying, idling nd eting in the fternoon. Significnt effects of ventiltion regimen x time (P < 0.05) were lso oserved for milk yield nd milk renneting prmeters, the LOV-30 ewes giving smller volumes of milk with deteriorted cogulting ehvior thn those of the MOV-60 group during the second hlf of the tril. When the renneting prmeters were gthered in the milk cogulting index (CoI), significnt differences (P < 0.05) were found etween the LOV-30 nd the MOV-60 group (Figure 3). No significnt differences emerged in ewe immune nd endocrine responses. Third tril. In exposed res, weekly verges of mximum mient tempertures were over 30 0 C throughout the study period nd often exceeded 35 C. Mximum mient tempertures nd reltive humidities were 3.9 to 6.8 C nd 6.3 to 12.3% lower in protected thn in exposed res. During dytime, verges of THI were lwys ner or over 80 in exposed res, wheres they only exceeded 75 during the second hlf of the tril. Solr rdition (P < 0.00l) nd the interction of solr rdition x time of feeding (P = 0.01) hd significnt effects on rectl tempertures (Figure 4). EXPM ewes hd higher rectl tempertures thn EXPA ewes, which in turn exhiited higher RT compred to PROM nd PROA ewes. EXP groups lso hd significntly higher respirtion rtes (P < 0.01) thn PRO groups. Immune response ws lower (P < 0.001) in EXPM ewes t d 10 nd in oth EXPM, EXPA nd PROM nimls (P < 0.05) t d 20 compred to PROA ewes (Figure 5). Exposure to solr rdition resulted in decresed plsm concentrtions (P < 0.05) of lnine minotrnsferse (ALT/GPT) (P < 0.05), lkline phosphtse (ALP) (P < 0.05), Potssium (P < 0.05), Mgnesium (P < 0.05) s well s in incresed levels of NEFA (P < 0.01) nd sprtte mino-trnsferse (AST/GOT) (P < 0.05). Milk yield nd composition were not chnged y exposure to solr rdition nd time of feeding, ut the EXPM tretment resulted in lower yields of csein nd ft (P < 0.05) nd reduced clot firmness (P < 0.05) compred with the three other tretments. Milk SCC ws similr cross tretments, ut PMNLC ws higher (P < 0.0l) in EXPM thn in PROM nd PROA milk. EXPM nimls lso hd the gretest mounts of totl nd fecl coliforms (P <0.0l) nd of Pseudomondcee (P < 0.05) s well s the highest numer of mstitis relted pthogens in their milk. Conclusions Our results suggest tht fn ventiltion system progrmmed to operte over upper criticl temperture nd humidity is not economiclly ttrctive in diry sheep housing. In fct, it involved out three fold greter energy cost nd did not led to remrkle improvements of ewe welfre nd productivity compred to n intermittent regimen split in 25 min/h ventiltion cycles during the wrmest hours of the dy. A further reduction of ventiltion cycles to 12.5 min/h resulted in ewe displying ltered ehviour, immune nd endocrine responses nd giving lower yields of milk. Therefore, our findings indicte tht ventiltion regimen, providing ventiltion cycles during the wrmest hours of the dy nd the night t men ventiltion rte of 66 m 3 /ewe per hour my dequtely sustin the welfre nd production performnce of lctting ewes rised in Mediterrnen climtes during summer. Douling ir speed from 2 to 4 m/s nd ventiltion rte from 35 to 70 m 3 /h per niml did not led to ny significnt improvement of ewe well-eing nd performnce in the groups sujected to 30 min ventiltion cycles. Insted, the ventiltion rte eing kept t 70 m 3 /h per niml, the group sujected to short ventiltion cycles displyed more intense ctivtion of thermoregultory mechnisms during the wrmest prt of the study period compred to the group enefiting from 60 min ventiltion cycles. Exposure to solr rdition under mient tempertures over 35 C prevented ewes from mintining their therml lnce. Provision of shde plyed mjor role in helping the
Interntionl Conference: Septemer 15-17, 2008 Rgus - Itly Innovtion Technology to Empower Sfety, Helth nd Welfre in Agriculture nd Agro-food Systems lctting ewe to minimize the dverse effects of high mient tempertures on therml lnce nd energy nd minerl metolism. Reduction of het lod during the wrmest hours of the dy through feed dministrtion in lte fternoon ws eneficil to ewes in minimizing the impct of therml stress on their immune function nd udder helth. Both solr rdition nd time of feeding hd only minor impct on ewe production performnce. References Adll E.B., Koty E.A., Johnson H.D. 1993. Physiologicl responses to het-induced hyperthermi of pregnnt nd lctting ewes. Smll Ruminnt Reserch, 11, 125-134. Amrl-Phillips D.M., McGillirg A.D., Linderg G.L., Veenhuizen J.J., Yng J.W. 1993. Effects of decresed vilility of glucose for diry cows. Journl of Diry Science, 76, 752-761. Beede D.K., Collier R.J. 1986. Potentil nutritionl strtegies for intensively mnged cttle during therml stress. Journl of Animl Science, 62, 543-554. Cost M.J.R.P., Siv R., Souz R.C. 1992. Effect of ir temperture nd humidity on ingestive ehviour of sheep. Interntionl Journl of Biometereology, 36, 218-222. Curtis S.E. 1983. Environmentl mngement in niml griculture. Iow Stte Press, USA. Hrtung J. 1994. The effect of irorne prticulte on livestock helth nd production. In: I. Ap Dewi, R.F.E. Axford, I.F.M. Mri, H. Omed, Pollution in livestock production systems. CAB Interntionl, UK, 55:69. Klstrup O., Nkken G., Brmley J., Bushnell R. 1987. Environmentl influences on ovine mstitis. IDF (Interntionl Diry Federtion Bullettin no. 217), Brussels, Belgium. Kume S., Kurihr M., Tkhshi S., Shit M., All T. 1987. Effect of hot environmentl temperture on mjor minerl lnce in lctting cows. Jpnese Journl of Zootechnicl Science, 58, 764-770. Rylnder R. 1986. Lung diseses cused y orgnic dusts in the frm environment. Americn Journl of Industril Medicine, 10, 221-227. Schneider P.L., Beede D.K., Wilcox C.J. 1988. Nycterohemerl ptterns of cid-se sttus, minerl concentrtions nd digestive function of lctting cows in nturl or chmer het stress environments. Journl of Animl Science, 66, 112-125.
Interntionl Conference: Septemer 15-17, 2008 Rgus - Itly Innovtion Technology to Empower Sfety, Helth nd Welfre in Agriculture nd Agro-food Systems 1.4 Antiody titer to OVA 1.2 1 0.8 0.6 0.4 0.2 0 11 21 30 40 Time (Dy) Low ventiltion rte (33 m3/h) Progrmmed ventiltion rte Moderte ventiltion rte (66 m3/h) Figure 1. Lest squre mens ± SEM of ntiody response to chicken egg lumin injection in ewes sujected to low (LVR), moderte (MVR) nd progrmmed ventiltion regimen (PROGR) 105 Cortisol (ng/ml) 85 65 45 25 Low ventiltion rte (33 m3/h) Moderte ventiltion rte (66 m3/h) Progrmmed ventiltion rte 5 0 60 120 240 Time (min) Figure 2. Lest squre mens ± SEM of plsm cortisol levels fter porcine ACTH injection in ewes sujected to low (LVR), moderte (MVR) nd progrmmed ventiltion regimen (PROGR) Milk cogulting index (CoI), mm/min 4.5 4 3.5 3 2.5 2 1.5 1 1 2 3 4 5 6 LOV-30 MOV-30 MOV-60 T ime (Weeks) Figure 3. Lest squres mens ± SE of cogulting index (CoI) in ewe milk s ffected y low ventiltion regimen (LOV-30), nd moderte ventiltion regimens providing short (MOV-30) nd long ventiltion cycles (MOV-60)
Interntionl Conference: Septemer 15-17, 2008 Rgus - Itly Innovtion Technology to Empower Sfety, Helth nd Welfre in Agriculture nd Agro-food Systems 41 40.8 40.6 Rectl temperture, C 40.4 40.2 40 39.8 39.6 39.4 39.2 39 c c 1 2 3 4 5 6 PROA PROM EXPA EXPM Weeks Figure 4. Rectl temperture of ewes when protected or exposed to solr rdition nd fed in the morning (PROM, EXPM) or in the fternoon (PROA, EXPA) Skinfold thickness (m m) 12 10 8 6 4 2 0 10 20 32 Dys PROA PROM EXPA EXPM Figure 5. Immune response to PHA injection in ewes when protected or exposed to solr rdition nd fed in the morning (PROM, EXPM) or in the fternoon (PROA, EXPA)