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Nepotpuna prehrana životinja pa tako i ovaca, može dovesti do narušavanje homeostaze organizma, smanjenja proizvodnje, pojave bolesti pa i do uginuća. Vitamin D označava skupinu usko povezanih tvari koje posjeduju antirahitičnu aktivnost. Među vitaminima jedinstven je po tome što je jedini koji se u organizam ne unosi samo hranom (sijeno, a u mesojeda hrana animalnog podrijetla), već se u organizmu (koži) stvara i uz pomoć sunčeve svjetlosti. Uobičajeno je da pašno držane ovce rijetko kad trebaju dodatke vitamina D. To je stoga jer ovce ukoliko su izložene UVB zračenju, odnosno sunčevoj svjetlosti mogu nadoknaditi manjak vitamina D u hrani sintezom vitamina D unutar kože. No, ukoliko je izloženost ovaca suncu smanjena dugotrajnom naoblakom ili uzgojem u zatvorenom te kada je sadržaj vitamina D2 u hrani nizak, tada unesena količina vitamina D može biti nedostatna. U janjadi se deficit vitamina D očituje pojavom rahitisa, a u odraslih ovaca pojavom osteomalacije te hipokalcemije. U ovaca se obično javlja u kasnom graviditetu, ali je zabilježena i u životinja različite dobi. Slijedom svega navedenoga cilj je ovoga preglednoga rada je upoznati doktore veterinarske medicine s novim saznanjima o ulogama i metabolizmu vitamina D u ovaca te njegovom značenju za zdravlje i održavanje proizvodnih sposobnosti jedinke, odnosno stada., The incomplete nutrition of animals, including sheep, can lead to a disruption of homeostasis, reduced production, development of disease, and even animal death. Vitamin D refers to a group of closely related substances that possess antirachitic activity. This is the only vitamin that is not only taken into the organism by way of dietary origin (hay, food of animal origin in carnivores), but is also synthetised in the body (skin) with the help of sunlight. Typically, grazing sheep rarely need vitamin D supplements. This is because sheep can compensate for the lack of vitamin D in their food by synthesizing vitamin D in the skin if they are exposed to UVB radiation or sunlight. However, if sheep exposure to the sun is reduced by prolonged bad weather or indoor animal breeding, and when the vitamin D2 content of the diet is low, then the animal may have insufficient vitamin D consumption. In lambs, vitamin D deficiency is manifested by rickets, and in adult sheep by osteomalacia and hypocalcaemia. In sheep, this usually occurs in late pregnancy but has been observed in animals of all ages. Following the above, the aim of this review is to acquaint veterinarians with new knowledge about the roles and metabolism of vitamin D in sheep and its importance for the health and maintenance of productive abilities of the individual animal or herd.

U ovom radu predstaviti će se jedinstvenost inovacije istraživanja provedenih u okviru HRZZ projekta “Inovativni funkcionalni proizvodi od janjećeg mesa“ (IP-2016-06-3685). Naime, uvjerenja smo da naša projektna inovacija, koja je prvenstveno rezultat znanja stečenog kroz proces istraživanja projekta, može predstavljati i instrument za povećanje konkurentnost i pokretanje razvoja poduzeća te tako pridonijeti gospodarskom i društvenom boljitku R. Hrvatske.

The adequate amounts of essential minerals are critical to maximizing the productivity and health of livestock. It is known that mineral deficiencies can lead to impaired growth and reproduction of animals and increases in disease, due to impaired immune function, and serious deficiencies may even cause death. Mushrooms are an excellent source of many nutrients including minerals, e.g. phosphorus, potassium and calcium but have a low sodium content. The objective of this study was to examine whether the intake of the dry and raw fruiting bodies of white button mushrooms does influence minerals serum concentration in lambs of the Lika Pramenka sheep breed. For this study, 60 clinically healthy, three-month-old lambs (32 females and 28 males), were randomly divided into four groups comprised of 15 lambs each. The first group was a control group in which lambs were fed for six weeks with a standard diet, the second group of lambs was fed for six weeks with commercial lamb food supplemented with 15% of raw Agaricus bisporus the third group was fed for six weeks with commercial lamb food supplemented with 1.5% dried Agaricus bisporus, and the forth group was kept extensively on pasture. Blood samples were taken once, and in the obtained sera calcium, potassium, chloride, sodium and phosphorus concentrations were defeminated by using standard commercial kits Beckman Coulter Biomedical Ltd., O’Callaghan’s Mills, Co. Clare, Ireland on an automatic Beckman Coulter AU 680 analyzer Beckman Coulter Biomedical Ltd, Ireland. Results showed that feed supplemented with Agaricus bisporus mushroom did not have a statistically significant effect on the selected blood minerals concentrations in lambs but also did not have any adverse effect on it. This research was funded by the Croatian Science Foundation with the project "Innovative functional lamb products" (IP-2016-06-3685).

Inovacije i kreativnost od iznimne su važnosti za gospodarski rast i razvoj duštva Hrvatske. U Europskoj Uniji pa tako i republici Hrvatskoj, trokut znanja između obrazovanja, istraživanja i inovacija čine temelj za postizanje sinergijskih učinaka ulaganja u znanost i inovacije iz nacionalnih, regionalnih i međunarodnih fondova. U ovom radu opisati ćemo jedinstvenost inovacije istraživanja provedenih u okviru HRZZ projekta “Inovativni funkcionalni proizvodi od janjećeg mesa“ (IP-2016-06- 3685). Rezultati projektnih istraživanja ukazuju da je janjad pasmine lička pramenka kojoj je u dnevni obrok bila umješana biomasa bijele i/ili smeđe plemenite pečurke bila dobrog zdravlja te se tom inovativnom tehnologijom uzgoja i hranjenja janjadi proizvela janjetina sniženog udjela kolesterola, povoljnog masnokiseliskog i aroma sastava te su sa pet inovativnih receptura proizvedeni funkcionalni polutrajni i trajni proizvodi od janječeg mesa.

Cilj ovog istraživanja bio je utvrditi učinak suhih i svježih ostataka peteljke plemenite pečurke (Agaricus bisporus) kao dodatka prehrani na oksidativno/antioksidativni status te razine adiponektina i leptina u krvnom serumu janjadi ličke pramenke. Cilj je također bio utvrditi postoji li učinak spola na istraživane pokazatelje u krvnom serumu janjadi. Ukupno 45 janjadi pasmine lička pramenka (3 mjeseca ; 22, 5±1, 7 kg tjelesne mase) nasumično je raspoređeno u tri skupine. Svaka se skupina sastojala od 15 životinja (8 ženki i 7 mužjaka) te je hranjena 6 tjedana. U I. skupini janjad je hranjena omercijalnom hranom kojoj je dodano 1, 5 % osušene plemenite pečurke. U II. skupini janjad je hranjena komercijalnom hranom s dodatkom 15 % svježe plemenite pečurke. Kontrolna skupina hranjena je standardnom (komercijalnom) prehranom. Uzorci krvi uzeti su jednokratno, a u dobivenom serumu su određeni pokazatelji oksidativno/antioksidativnog statusa i adipocitokini: lutation peroksidaza (GPX), superoksid dismutaza (SOD), malondialdehid (MDA), leptin i adiponektin. Vrijednosti GPX-a, SOD-a, leptina i adiponektina određene su ELISA-om. Koncentracija MDA određena je tekućinskom kromatografijom visoke učinkovitosti. Niti jedan od istraživanih pokazatelja nije se značajno razlikovao u serumskim vrijednostima između skupina janjadi oba spola. No, kada se uključio utjecaj spola, janjice u skupini II. imale su značajno višu vrijednost MDA u odnosu na janjice u skupini I. Značajno više vrijednosti SOD-a i adiponektina određene su u kontrolnoj skupini janjica u odnosu na janjice u skupini II. Prema dobivenim rezultatima može se zaključiti da je dodatak prehrani od 1, 5 % osušene plemenite pečurke imao povoljniji učinak na oksidativno/antioksidativni status i vrijednosti adiponektina u krvnom serumu ženske janjadi u odnosu na žensku janjad hranjenu s dodatkom od 15 % svježe plemenite pečurke. Dodatak prehrani od osušenih i svježih pripravaka plemenite pečurke nije narušio homeostazu oksidativno/antioksidativnog statusa i vrijednosti adipocitokina u krvnom serumu janjadi.

Blood profiles, which includes haematology parameters provide a useful tool to assess the physiological status of reptiles that may be influenced by several internal and environmental factors. In the available literature, data on blood morphometric parameters in snakes are not available or are very scarce. Therefore, the aim of current study was to determine the values of red blood cell (RBC) morphometric size and shape parameters and to investigate the differences in RBC morphometric parameters between genders in corn snakes (Pantherophis guttatus). Blood samples, from 7 snakes (3 males ; 4 females), were taken from the ventral tail vein. The smears were confectioned on slides immediately after the blood collection and stained according to the May-Gründwald Giemsa method. Morphometric analyses were performed on 700 RBCs in stained blood smears using SFORM computer-assisted program. Following morphometric RBCs and RBCs nucleus size, parameters were determined: area, outline, convex area, minimal and maximal radius, length and breadth, as well as calculated morphometric RBCs shape parameters: ellipticity, elongation, solidity, roundness, form factor, contour index. Significantly higher values of RBCs maximal radius, length, ellipticity and elongation were established in females, respectively. Conversely, significantly higher values of RBCs nucleus maximal radius and length were determined in male snakes. In conclusion, males had significantly wider erythrocytes with a longer nucleus, and females, on the contrary, more elongated, elliptical erythrocytes with a wider nucleus. Knowledge of physiological morphometric parameters of snake RBC and physiological differences within the species is a prerequisite for the correct interpretation of pathologically altered erythrocyte morphology.

The data on semi-aquatic turtles’ erythrocytes morphometric parameters still are not known, so the aims of this study were to determine the values of erythrocytes morphometric size and shape parameters and their differences between four pet turtle’s species. Blood samples of four turtle’s species: Trachemys scripta elegans (TSE ; n=5), Trachemys scripta scripta (TSS ; n=2), Graptemys pseudogeographica kohni (GPK ; n=2), Sternotherus odoratus (SO ; n=2) were taken from the dorsal coccygeal vein. Smears were stained according to the May-Gründwald Giemsa method. By using a computer assisted-image analyses system, morphometric measurements were performed (100 erythrocytes per smear) with the "SFORM" program. The size parameters of erythrocytes and erythrocytes nucleus (area, outline, convex area, minimal and maximal radius, length and breadth), as well as shape parameters for erythrocytes (ellipticity, elongation, solidity, roundness, form factor and contour index) were determined. In the TSS turtles significantly higher values of erythrocytes area, convex area, minimal and maximal radius and length were determined, respectively. Also, in the TSS turtles, significantly higher values of erythrocytes nucleus area, outline, convex area, minimal and maximal radius, length and breadth were found. According to the obtained data, we can conclude that the morphometric parameters of erythrocytes differ between turtle species, i.e. the TSS turtles have the largest erythrocytes with the largest nucleus. The findings in this study can be used as the baseline values in the assessment of health of the above species and also will be useful in future haematological studies related to these species.

To the best of our knowledge, this is the first analysis of morphometric parameters for assessing the influence of radiofrequency radiation (RF) at 2500 MHz on spermatozoa subpopulations. The aim of this study was to establish the influence of RF at 2500 MHz in vitro exposed breading boars’ semen on proportions of spermatozoa subpopulations using the principal component (PC) and cluster analysis according to morphometric head and tail parameters. Semen of 12 breeding boars was collected using a manual fixation method. Semen samples of each boar were divided into a control and an experimental sample. The experimental samples were exposed to RF of 2500 MHz in gigahertz transverse electromagnetic chamber for 2 hours at an electromagnetic field strength of 10 V/m. Semen smears were stained using the Spermac method. Morphometric analyses in the control and experimental samples were performed in stained smears by SFORM computer- assisted program. Morphometric size and shape parameters of boar’s spermatozoa (n=1691) were determined. The PC analysis revealed four components with the most important value for each (head length and width, head regularity, midpiece outline). Cluster analysis indicated there were two subpopulations (S1 - spermatozoa of smaller length and larger head width, more regular head shape and smaller circumference of the midpiece and S2 - spermatozoa of longer length and smaller head width, more elongated head shape and larger outline of the midpiece). Exposed boars’ semen samples had a greater proportion of S1 spermatozoa compared to control samples (51.83% vs 48.7%) and a lesser proportion of S2 spermatozoa (47.54% vs 52.47%). The effect of RF at 2500 MHz in vitro exposed breading boars’ semen was manifested in a reduced proportion of spermatozoa subpopulation with the spermatozoa of more elongated head and larger midpiece outline which could decrease spermatozoa motility and fertility potential.

Mushrooms contain a variety of active substances and, among others, could provide functional organic selenium (Se). Se is an incorporated element into selenoprotein glutathione peroxidase (GSH-Px), a significant factor in protecting cells from oxidative stress. GSH-Px measured in blood adequately reflect their level in other tissues and systemic oxidative status. The aim of this study was to determine whether intake of the dry and raw fruiting bodies of white button mushrooms influences blood serum oxidative status in Lika pramenka lambs. The study was conducted on 45 lambs of the Lika pramenka sheep breed at the age of three months in the Crkvina, Karlovac County, Croatia. The lambs were randomly divided into three groups, with 15 animals in each group (8 females and 7 males). Each group was fed for 6 weeks: control (basal diet), experimental I. (basal diet with 15 % of raw Agaricus bisporus) and experimental II. (basal diet with 1.5 % dried Agaricus bisporus). Blood was collected from the jugular vein using vacuum tubes without anticoagulants. The activities of GSH-Px, superoxide dismutase (SOD), and the concentration of malondialdehyde (MDA) were determined in the obtained serum. The differences in oxidative status between groups were not statistically significant, although there were slightly higher serum GSH-Px and MDA values found in lambs fed with 15% of raw white button mushroom supplements. It can be concluded that white button mushroom supplementation does not affect the lamb’s serum oxidative status. Acknowledgement: This study has been fully supported by the Croatian Science Foundation under the project “Innovative functional lamb meat products (“Inojanjetina”)” (HRZZ‐IP‐06‐ 2016‐3685).

In the available literature, the immunostimulatory effect of the button mushroom (Agaricus bisporus) dietary supplement mixed in the daily rations has been described in monogastric animals kept in controlled environment and intensive production conditions. Thus, after 42 days of feeding with the Agaricus bisporus dietary supplement, an increase in the proportion of T and B lymphocytes was recorded in the peripheral blood of domestic pigs and broilers. The aim of the research was to determine the effect of Agaricus bisporus on the kinetics and change in the proportion of memory T cells in lambs. Thirty lambs of the Lika pramenka breed at the age of three months were used in the experiment. The lambs were divided into two groups: control (without addition of Agaricus bisporus supplement in daily rations) and experimental (with addition of 1.5% dry Agaricus bisporus supplement in daily rations). On days 0 and 42 of the experiment, cytometric analysis of lamb peripheral blood lymphocytes was performed using directly fluorochrome- labeled specific mouse monoclonal antibodies for memory T cells of sheep (antibody panel: CD45+ CD3+ CD8+ CD4+). Statistical data analysis was performed using the SAS 9.4. software package. On the 42nd day of the experiment, in lambs fed with the addition of 1.5% dry Agaricus bisporus supplement in daily rations, a 0.33% higher share of memory T cells was recorded compared to control group lambs. There is a general lack of scientific information about the immunomodulatory effect of Agaricus bisporus in ruminants. Therefore, the results obtained in this paper are scientific recommendations for further assessment of T cells kinetics, as well as other subpopulations, during and after feeding with Agaricus bisporus dietary supplement. This study has been fully supported by the Croatian Science Foundation under the project “Innovative functional lamb meat products” (HRZZ-IP-06-2016-3685).

Morfometrijske analize eritrocita, utvrđene pomoću sofisticiranih i naprednih računalnih programa su točnije i preciznije od konvencionalnih, mikroskopskih analiza. Automatizirana metodologija morfometrijske analize oblika eritrocita omogućuje kvantificiranje oku nevidljivih promjena morfometrijskih značajki, te boljem razumijevanju kliničkog utjecaja istih promjena na organizam. Do sada još nisu istraživane morfometrijske osobitosti veličine i oblika eritrocita u pasa s piroplazmozom. Stoga, ovi rezultati ujedno prikazuju promjene morfometrijskih pokazatelja na samom eritrocitu, no također otvaraju vrata kompjuterskoj automatizaciji dijagnostike piroplazmoze prepoznavanjem specifičnih odstupanja od fizioloških raspona morfometrijskih pokazatelja. Istraživanje je provedeno na dvanaest standardno obojenih krvnih razmaza pasa pozitivnih na piroplazmozu prije aplikacije terapije. Pomoću svjetlosnog mikroskopa pri povećanju od 1000x uz korištenje imerzione tekućine, kamerom visoke rezolucije u boji slikani su eritrociti s inkluzijama Babesiae canis canis te eritrociti bez inkluzija. Ukupno je slikan tristo osamdeset i jedan eritrocit. Izmjereni su slijedeći jednostavni morfometrijski pokazatelji veličine eritrocita: duljina, širina, najmanji i najveći polumjer, površina, konveksna površina ili ispupčenost i opseg, te su izračunati slijedeći morfometrijski pokazatelji oblika eritrocita: eliptičnost, izduljenost, zaobljenost, faktor zaokruženosti oblika, indeks ispunjenosti i indeks pravilnosti površine eritrocita. Statistička analiza odrađena je pomoću programskog paketa SAS 9.4. Hipoteza ovoga rada temeljena je na pretpostavci da prisutnost inkluzije unutar eritrocita dovodi do određenih promjena njegove građe i oblika. Cilj rada bio je kvantificirani promjene morfometrijskih pokazatelja eritrocita s inkluzijama Babesiae canis canis, ali i odrediti postoji li korelacija broja inkluzija te razvojnog stadija sa stupnjem deformacije membrane. Interpretacijom dobivenih rezultata procijenjena je i funkcionalnost eritrocita u svakom pojedinom stadiju kliničkog manifestiranja piroplazmoze. Rezultati dobiveni morfometrijskim mjerenjem invadiranih eritrocita ukazivali su da se membrana eritrocita do trenutka rupture tj. hemolize deformira. U usporedbi neinvadiranih i invadiranih eritrocita zaraženih pasa očitovala se statistički značajna razlika u gotovo svim morfometrijskim pokazateljima veličine i oblika eritrocita. Morfometrijskim mjerenjem eritrocita invadiranih Babesiom canis canis bilo je vidljivo da su eritrociti postali eliptičniji, izduljeniji s većim indeksom ispunjenosti i manjom zaobljenošću. Dakle, prisutnošću bilo kojeg broja i bilo kojeg razvojnog stadija protozoe dolazi do deformacije membrane te eritrocit gubi fiziološki diskoidalni oblik. Gubitak fiziološkog oblika eritrocita i povećanje ukupnog volumena utječe na njegovu mobilnost kroz periferne kapilare što pogoduje razvoju kompliciranog oblika piroplazmoze. Također su bile vidljive i značajne varijacije između pasa u morfometrijskim pokazateljima. Psi koji su uzorkovani unutar 24 sata od pojave prvih simptoma ili pronalaska krpelja imali su slabije izražene deformacije membrane. Do dostizanja kritičnog hemolitičkog volumena funkcija eritrocita je vjerojatno djelomično očuvana. Ovi rezultati dodatno naglašavaju važnost pravovremenog prepoznavanja bolesti i započinjanja prikladne terapije kako bi se smanjila smrtnost kod piroplazmoze.

Serum biochemical parameters are used to evaluate animal condition and detect diseases. The gender-related differences in physiological processes, pathophysiology, diagnosis and treatment of diseases have an increasing importance in human medicine, and these differences should be investigated and implemented in animal models. Therefore, the aim of this study was to investigate the effects of gender and gonadectomy in pigs on some blood serum biochemical parameters. The study was conducted on 20 clinically healthy, sexually mature pigs of the Swedish Landrace breed. The pigs were divided into four groups (n = 5 each): intact boars (age 271 ± 4 days), intact gilts (age 270 ± 4 days), castrated boars (age 267 ± 5 days) and castrated gilts (age 269 ± 4 days). Blood samples were taken from the jugular vein by using blood test tubes without anticoagulant. After centrifugation, the serum was separated, and the activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT), and γ-glutamyl transferase (GGT) as well as the concentrations of urea and glucose were determined spectrophotometrically. Data analysis was performed by using the Statistica v. 13.4 software. The normality of dana distribution was tested by using the Kolmogorov-Smirnov test, and the significance of the differences between the groups by using an analysis of variance and the Tukey HSD test. The study results showed that the castrated boars had a significantly higher (p < 0.05) urea concentration in the blood serum than the intact boars. In addition, the castrated boars had a significantly lower (p < 0.05) serum glucose concentration than the intact boars and gilts. The serum activities of AST, ALT and GGT were the highest in the castrated boars, and the lowest in the intact boars, except for the GGT activity, which was the lowest in the castrated gilts. It can be concluded that for the interpretation of serum biochemical parameters in pigs, i.e. to ensure an accurate clinical diagnosis and prognosis, gender and gonadectomy should be considered. However, further studies including a higher number of animals are required in order to define the effects of gender and gonadectomy on the values of biochemical parameters in blood serum of pigs more precisely.

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The knowledge that spermatozoa differ in morphological and functional characteristics has initiated studies in order to isolate subpopulation of spermatozoa with better functional abilities. The aim of this study was to determine spermatozoa subpopulations based on morphometric parameters of spermatozoa head (MPSH) and tail (MPST) in ejaculates of bucks (N=12) over a period of 3 months. Semen smears were prepared and stained with method of Bloom to establish the proportion of live spermatozoa. By using a computer assisted-image analyses system, morphometric measurements were performed (50 spermatozoa per smear) with the "SFORM" program. Morphometric data were collected for spermatozoa head, mid-piece and tail (area, outline, length, width), and for head shape (regularity, rugosity, ellipticity, elongation). By analyzing of principal components, two values were obtained explaining 85% of variance for MPSH and 78.5% for MPST. For MPSH, the 1st component focused on measurements of size and the 2nd on head shape. The 1st component for MPST focused on length and the 2nd on the width of the mid-piece. Cluster analysis of the most important parameters for each component revealed 4 spermatozoa subpopulations based on MPSH and MPST. The 1st subpopulation of MPSH comprised small sized spermatozoa with high rugosity spermatozoa (17%), the 2nd the biggest sized (4.8%), the 3rd elliptical and elongated (24.5%) and the 4th average sized spermatozoa (53.4%). The 1st spermatozoa subpopulation based on MPST comprised spermatozoa with the smallest size of the mid-piece (63.0%), 2nd spermatozoa with the widest mid-piece but the shortest tail (10.9%), the 3rd spermatozoa with the longest tail (21.6%) and 4th spermatozoa with the greatest length, area and outline of mid-piece (4.5%). Morphometric analysis of the principal components and cluster analysis, could provide data about semen quality and the freezing ability, according to the proportions of the individual subpopulations in the semen.

Morphological and functional characteristics of spermatozoa (SP) in the ejaculate of bucks depending on genetic and environmental factors. The assessment of SP morphology is more sophisticated today with the use of systems for computer-assisted morphometric image analysis. Such methods provide more precise and detailed measurements compared to standard subjective methods. The aim of this study was to investigate the influence of the age of bucks on the morphometric parameters of SP head and tail, and the association of SP tail morphometric parameters with the proportion of motile SP. During the experimental period of 3 months, the ejaculates of French alpine bucks (n=12), aged from 1-4 years, were collected weekly by artificial vagina. The motility of SP was determined in semen samples by microscopy. Morphometric analysis was performed on stained semen smears (n=144) using method of Bloom, by the computer assisted-image analyses system. The parameters of SP head and tail size such as: area, outline, length, width, and parameters for head shape: regularity, rugosity, ellipticity, elongation were determined. The bucks were categorized according to their age and the proportion of motile SP in ejaculate, into 3 groups for each category. The youngest bucks (1-1.5 years) had significantly greater outline and length of mid-piece and significantly greater head ellipticity and elongation compared to the other 2 groups (1.5-2.5, 2.5-4 years). The oldest bucks (2.5-4 years) had a significantly smaller elongation and greater rugosity of the head. Bucks with the highest proportion of motile SP in the ejaculate (≥85%) had a significantly greater outline, length and area of the mid-piece and length of tail compared to other 2 groups (75% ; 75-85% motile SP in the ejaculate). The obtained results suggest that the SP of younger bucks had functionally desirable morphometric characteristics, and that the bucks with larger proportion of motile SP in their ejaculate had larger mid-piece and a longer tail of spermatozoa.

Na ukupno 30 razmaza suspenzija izoliranih mononuklearnih stanica izmjerene su morfometrijske karakteristike limfocita inkubiranih 48 h u staničnom mediju bez ili uz dodatak konkanavalina A (Con A). Digitalna analiza krvnih stanica napravljena je na osobnom računalu s računalnim programskim paketom SFORM (VAMSTEC, Zagreb). Obradom rezultata utvrđene su značajno veće vrijednosti površine, opsega, maksimalnog polumjera, konveksne površine te faktora pravilnosti limfocita stimuliranih s Con A u usporedbi s istim pokazateljima stanice nestimuliranih limfocita. Vrijednosti minimalnog polumjera i dužine stanice nakon inkubacije bile su podjednake u oba inkubacijska medija. Mjerenjem morfometrijskih pokazatelja jezgara, nisu ustanovljene značajne promjene njihove površine ili opsega, međutim utvrđen je značajno smanjen minimalni polumjer i širina jezgre, značajno manji nukleo-citoplazmatski omjer te smanjena vrijednost faktora pravilnosti u limfocitima stimuliranim s Con A. Utvrđene promjene morfoloških pokazatelja stanica limfocita i njihovih jezgara upućuju na povećanu metaboličku aktivnost limfocita tijekom aktivacije s mitogenom te smatramo da morfometrijska analiza može pružiti vrijedne podatke u procjeni transformacije imunokompetentnih stanica.

Istražen je utjecaj dodatka organskoga selena u hranu na metabolizam masnih tvari, intenzitet lipidne peroksidacije te aktivnost antioksidacijskih enzima u srčanom mišiću pilića po završetku tova i stresa potaknutog 48-satnim gladovanjem. Istraživanje je provedeno na pilićima hibridne linije Ross 308 podijeljenim u dvije skupine: kontrolnu skupinu hranjenu tijekom uzgoja standardnim krmnim smjesama s 0, 15 ppm anorganskoga selena i pokusnu skupinu pilića kojoj je u standardne krmne smjese dodano 0, 3 ppm organskoga selena. Nakon završetka uzgoja pilića te nakon 48-satnog gladovanja žrtvovano je po 10 pilića iz kontrolne i po 10 pilića iz pokusne skupine. Nakon žrtvovanja životinja uzeti su uzorci srčanoga mišića u kojima su određene koncentracije triacilglicerola, kolesterola i reaktivnih spojeva tiobarbiturne kiseline te aktivnosti enzima lipoproteinske lipaze, glutation peroksidaze (GSH-Px), katalaze (CAT), bakar, cink superoksid dismutaze i manganske superoksid dismutaze (Mn-SOD). Nakon gladovanja aktivnost CAT-a (P

U radu su istražene spolne razlike i utjecaj gonadektomije (kastracije i ovarijektomije) na količinu i sastav masti, intenzitet lipidne peroksidacije, aktivnost antioksidacijskih enzima te kakvoću mesa u smislu tehnoloških svojstava i kemijskog sastava u uzorcima velikog slabinskog mišića (m. psoas major) svinja po završetku tova. Istraživanje je provedeno na tovnim svinjama pasmine švedski landras. U dobi od četiri tjedna, obavljena je kastracija (5 mužjaka) i ovarijektomija (5 ženki), te lažna kastracija (5 mužjaka) i lažna ovarijektomija (5 ženki). Na uzorcima mesa određene su aktivnosti glutation peroksidaze, superoksid dismutaze, gama glutamil transferaze i laktat dehidrogenaze te koncentracije kolesterola, triacilglicerola, fofsolipida i malondialdehida. Nadalje, utvrđen je utjecaj spola i kastracije odnosno ovarijektomije na kakvoću mesa u smislu njegovih tehnoloških svojstava i kemijskog sastava. Prema dobivenim rezultatima utvrđene su značajne spolne razlike količine i sastava masti. Spol svinja nije imao značajniji učinak na oksidativnu stabilnost mišića. Vezano uz gonadektomiju rezultati upućuju da je veliki slabinski mišić ovarijektomiranih ženki najosijetljiviji na oksidacijske promjene. Prema kriterijima Hofmann-a (1994.) i Van Laack-a (2000.), na osnovi rezultata za pH, uzorke mesa ovarijektomiranih ženki možemo smatrati BMV mesom. Analize udjela masti ukazuju na značajnu razliku uzoraka ovarijektomiranih ženki u odnosu na ostale pokusne skupine.

Cilj ovog istraživanja je detaljan opis i određivanje morfometrijskih pokazatelja krvnih stanica komarče, te njihova usporedba s obzirom na dob. Na ukupno 24 krvna razmaza mlađi u dobi od osam mjeseci te konzumnih komarči u dobi od dvije godine prepoznati su i opisani eritrociti i trombociti, te pet vrsta leukocita: limfociti, monociti, acidofilni granulociti 1 i 2 te neutrofilni granulociti. Morfološke razlike i analiza morfometrijskih pokazatelja ukazuju na to da su acidofilni granulociti 1 i 2 ustvari dvije različite vrste granulocita: eozinofili i heterofili.

Marek’s disease is a common lymphoproliferative disease of chickens, usually characterized by mononuclear cell infiltration of peripheral nerves and various other organs and tissues, including the iris and skin. A group of 70 newly-hatched chicks was vaccinated by means of nebulization and exposed for 60 seconds to the HVT FC 126 vaccine, while the other group of 70 chicks received the same vaccine by parenteral route (s.c.). The aim of this study was to compare the morphological patterns of the chicken’s lymphocytes in their peripheral blood, before and after vaccination by means of either nebulization or parenteral injection. Image analysis was performed using the SFORM software (VAMSTEC, Zagreb, Croatia). A total of 50 blood smears from vaccinated chickens (20 by means of nebulization and 30 by parenteral injection), with an average of 100 cells per smear, were analyzed. The results showed that the peripheral blood lymphocytes of chickens vaccinated by means of nebulization, compared with parenteral vaccinated, had significantly higher values for the majority of the measured variables: area, outline, minimum and maximum radius, length, breadth and convex area, on all days after vaccination, except on day 4, indicating the significantly higher metabolic activity of those cells. In lymphocytes of chickens vaccinated parenterally, only the nucleo-cytoplasmic ratio was higher. The results show that morphological patterns of immunocompetent cell transformation could be used to evaluate immune responses to vaccination and vaccine efficacy. We conclude that nebulization as a mode of vaccination against Marek’s disease stimulates the transformation of immunocompetent cells much earlier, thus shortening the time of immunosuppression and improving the immune response. This is of paramount importance for the practical application of Marek’s disease vaccine.

Housing conditions of farming animals often act as stressing factors decreasing the efficiency of animal production. Active components of many plants have evolved as antibacterial and antioxidant agents against environmental stress due to a variety of oxidizing and potential harmful free radicals. Adaptive mechanisms of cells to stres conditions lead to changes, increase and/or abnormalities of cells and nuclei, that can be quantified by morphometry. Morphometric parameters of erythrocytes, lymphocytes and monocytes were measured on the 21st and on the 42nd day in the control group, and in experimental group of chicken fattening with essential oils Aroma korm® and clinoptilolite as food additives. On the total of 36 blood smears area, outline, minimum and maximum radius, convexity, length and breadth of the whole cells and their nuclei as well as nucleo/cytoplasmic ratio were measured. Digital image analysis was performed using the SFORM software (VAMSTEC, Zagreb, Croatia). On the 21st day of fattening with essential oils as food additives almost all measured erythrocyte parameters were lower in comparison with the control group, while on the 42nd day values increased and exceeded the control values. Increased size and irregular shape of lymphocytes and their nuclei were seen in experimental group on the 21st day, but on the 42nd day all measured parameters were reduced in comparison to the values of control group. The size of monocytes did not change until the 42nd day, when they had significantly smaller outline and convexity. Established morphological changes of erythrocytes indicate, after the initial stress reaction, more intensive erythropoiesis in experimental chickens. Reduced size and convexity of lymphocytes and monocytes on the 42nd day in experimental groups of chickens were in the accordance with anti-inflammatory and antimicrobial effects of fennel and lemon essential oils, as well as zeolite effectiveness in reducing total flora contamination on the intestine. Obtained results confirmed that morphometric analysis can provide valuable data for assessing blood cell adaptive responses and could complement conventional hematology.

U ovom znanstvenom preglednom radu opisuju se endotoksini u zraku nastambi za perad, njihove prosječne koncentracije, način određivanja te utjecaj na zdravlje životinja i ljudi.

IMUNO-2865® is a natural hemicellulose compound with well-documented immunostimulative properties in people and marine mammals , but not yet tested on teleost fish. The aim of this study was to test in vivo effects of IMUNO- 2865® on blood biochemistry parameters in sea bream under aquaculture conditions during winter period. Blood Ca++ levels were significantly higher after 60 days of feeding in all treatment groups compared to the control, and remained elevated in group 4 even after 90 days following secession of supplementation. Antioxidant enzyme (SOD) was increased in group 4 compared to the control group from day 60 until the end of the experiment (P

Brzi rast pilića u tovu prati smanjena aktivnost te produženi period sjedenja i ležanja što s obzirom na gustoću smještaja i njen utjecaj na kvalitetu stelje predstavlja glavni čimbenik smanjenja dobrobiti. Visoka koncentracija amonijaka u vlažnoj stelji utječe na pojavu kontaktnog dermatitisa u području tarzusa i prstiju. U ovom preglednom radu opisane su dosadašnje spoznaje o utjecaju gustoće smještaja i obogaćenja okoliša na pojavu kontaktnog dermatitisa tarzusa i prstiju te procjena dobrobiti analizom morfoloških svojstava autopodija pilića u tovu. Planira se provesti istraživanje, koje će uključiti obogaćenje okoliša, uz pretpostavku veće mišićne aktivnosti pokusne skupine pilića u tovu u odnosu na konvencionalno držane te očekivanu razliku u broju i veličini svih mišićnih vlakana pilića. Istražit će se morfološka svojstva mišića s obzirom na različite gustoće naseljenosti, građu bedrene kosti i goljenice (oblik i veličina kosti) te odnos debljine kompaktne kosti i medularne šupljine kosti. Također će biti istražena morfološka svojstva bedrenog i prsnoga mišićja s obzirom na svojstva mišićnih vlakana.

Aquaculture is one of the worlds fastest growing food industries. There is, however, an over- dependence on the use of prophylactic and therapeutic antibiotics in aquaculture, with a significant cost to human and environmental health (bacterial resistance, residues, toxicity). Alternative products are being evaluated worldwide in the search for natural and cost-effective prophylactic supplements that can be incorporated into the fish diet. IMUNO-2865® is a natural hemicellulose compound with well-documented immunostimulative properties in people, terrestrial and marine mammals. The aim of this study was to test in vivo effects of IMUNO-2865® on blood parameters and oxidative stress responses in sea bream under aquaculture conditions. The research was conducted over a period of 6 months at a sea bream farm in Central Adriatic (Croatia). A total of 640 sea bream specimens (18 month old, weighing 277.8± 38.0 g) were divided into four groups, placed in four 8m3 net cages and conditioned for two weeks. Fish were fed with commercial pelleted food (Skretting) at a rate of 10 g dry diet/kg biomass/day. Four experimental diets containing 0 g (control Group 1), 1g (Group 2), 10g (Group 3) and 25g (Group 4) of IMUNO- 2865®/kg feed were prepared ahead of time and stored at 4oC. Samples were collected on days 0, 30, 60 and 90, after which the regular feeding regime was resumed at 15g dry diet kg-1 biomass. A final sampling was performed on day 180 (90 days after consumption of the last experimental diet). No mortality was observed throughout the experiment. Although blood parameters fluctuated among sampling periods (e.g. erythrocyte counts were highest at the end of the experiment), no significant differences were noted among treatment groups for total number of erythrocytes, leukocytes, basophils, eosinophils, lymphocytes and thrombocytes. However, a significant increase (P

Abortion in cows represents a serious burden for dairy industry with a great economic impact. Most abortions are due to infectious diseases, several of them are zoonoses. Bacterial zoonoses such as leptospirosis, Q fever and brucellosis are important cattle infections associated with intrauterine death and reduced fertility. The aim of this study was to explore differential proteome profile in serum of cows that aborted due to subclinical infection of leptospirosis and Q fever in order to uncover the pathophysiological molecular mechanisms involved in abortion caused by infectious diseases.The study was conducted on 15 dairy cows divided into three groups: Group I (control, healthy cows, n=5) ; Group II (cows infected with Coxiella, n=5) and Group III (cows infected with Leptospira, n=5). Group II and Group III comprised cows that have aborted. In collected sera, 1D- and 2D-electrophoresis were carried out and the mass spectra were acquired with an Ultraflex III MALDI-TOF/TOF spectrometer (Bruker-Daltonics).Six proteins were found to be (p≤0.05) either up-regulated or down-regulated in bovine sera with Q fever and leptospirosis in comparison to the control. This homology between the two pathologies might suggest that they share some common features that can cause abortion.

Psoas major muscles had a high percentage of slow-twitch oxidative fibres, numerous mitochondria, and possess a high capacity for oxidative metabolism. Since susceptibility of pig muscle to lipid oxidation depends upon relative proportion of fibre type, amount and composition of lipids, and prooksidant/antioxidant balance, the aim of this study was to investigate sex differences of antioxidant enzymes activities, lipid content and lipid peroxidation in swine’s psoas major muscle. The experiment was carried out on twenty Swedish Landrace pigs reared under intensive management system. At four weeks of age five males and five females were gonadectomized, and the rest of the pigs were sham operated in the same way without testes and ovary removal (control males, n = 5 ; control females n = 5). Clinically healthy pigs were slaughtered at ten months of age. In the collected and homogenized muscles samples glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), triacylglycerides, phospholipids and cholesterol, as well as end product of lipid peroxidation (malondialdehyde, MDA) were determined. The results showed no differences between intact males and females antioxidant enzymes activities and MDA concentration, but female pigs had significantly higher cholesterol and lower triacylglycerides concentrations than males. Barrows had a significantly lower triacylglycerides concentration than control males, and castrated females had significantly higher triacylglycerides and phospholipids concentrations than barrows, as well as a significantly lower GSH-Px activity than control females and barrows. The results indicated that muscles of ovariectomized females are more susceptible to lipid peroxidation, due to the lowest GSH-Px activity, higher phospholipids content, and, although not significantly, the highest MDA concentration.

U ovom istraživanju izmjerene su promjene morfologije stanica i jezgri eritrocita, limfocita i monocita kontrolne skupine pilića, te 21. i 42. dana tova u skupini kojoj su u hranu dodana eterična ulja XTRACTᵀᴹ i prirodni zeolit. Morfometrija stanica krvi napravljena je na osobnom računalu s podržavajućim programom “SFORM” tvrtke VAMSTEC, Zagreb, te je analizirano ukupno 27 krvnih razmaza. Statistička obrada rezultata izvršena je uz pomoć kompjuterskog programa za analizu podataka STATISTIKA, 12. Digitalna analiza morfometrijskih pokazatelja pokazala je da se veličina eritrocita značajno smanjila 21. dana u odnosu na kontrolu, dok su 42. dana vrijednosti ponovno porasle na razinu kontrolne skupine. Zabilježena je povećana nepravilnost oblika i povećanje veličine limfocita 21. dana, što je tijekom 42. dana ostalo konstantno povećano. Povećanje veličine i nepravilnosti oblika monocita nastupio je tek 42. dana. Rezultati analize morfometrijskih pokazatelja eritrocita, leukocita i monocita potvrdili su njen značaj u procjeni adaptivnih odgovora krvnih stanica na stresne čimbenike tijekom intenzivnog tova. Rezultati također potvrđuju imunomodulacijski učinak eteričnih ulja dodanih u hranu pilića, što se očitovalo statistički značajnim razlikama nađenim u svim istraživanim varijablama stanica i jezgri limfocita i monocita: površini, opsegu, minimalnom i maksimalnom radijusu, zakrivljenosti, dužini i širini. Postignuti rezultati mogu biti od koristi u promicanju učinkovitih metoda za procjenu imunokompetencije organizma te dijagnosticiranje i prevenciju stresnih stanja pri intenzivnom tovu pilića.

The aim of this research was to investigate the effects of essential oils and natural zeolite on blood cells morphology of chickens. Cells and nuclei morphometric parameters of erythrocytes, lymphocytes and monocytes were measured on the 21st and on the 42nd day in the control group, and in chicken fattening with essential oils XTRACTᵀᴹ and clinoptilolite as food additives. On the total of 36 blood smears area, outline, minimal and maximal radius, convexity, length and breadth of the whole cells and their nuclei as well as nucleo/cytoplasmic ratio were measured. Digital image analysis was performed using the SFORM software (VAMSTEC, Zagreb, Croatia). On the 21st day the size of erythrocytes was significantly lower compared to the control group, but on the 42nd day values returned back to control levels. Increased size and irregular shape of lymphocytes were seen on the 21st day of fattening and remained the same on the 42nd day. The size of monocytes did not change until the 42nd day, when they were significantly larger compared to the control group. Obtained results of morphometric characteristics analysis confirmed its importance in evaluation of blood cells’ adaptive responses to stress factors during intensive fattening. Established morphological changes of examined variables of monocytes and lymphocytes imply to their transformation and indicate immunomodulatory effects of added essential oils. Given results can be used to promote effective methods for assessment of immune competence of the organism so far as the diagnosis and prevention of stress condition in broiler chickens

Cell proteins as well as unsaturated fatty acids are susceptible target for a free radical attack. As sex hormones exert a long-term regulation on tissue metabolism, particularly on synthesis, remodeling and utilization of amino acids and fatty acids, it was of interest to investigate the effect of castration on the activities of antioxidant enzymes, lipid peroxidation and protein carbonyl contents in the pheasant capon tissues. Thirty pheasants reared in a commercial pheasantry were included in the experiment. Half the pheasants were castrated at 8 weeks of age and the other half underwent sham surgery. In the 32nd week of age the pheasants were stunned, slaughtered, bled and pluck in a commercial slaughterhouse. In the tissue samples (heart, liver, duodenum and spleen) of intact and castrated animals, the activities of glutathione peroxidase (GSH-Px), total superoxide dismutase (TSOD), manganese superoxide dismutase (Mn-SOD), cooper, zinc superoxid dismutase (CuZn-SOD), glutathione, thiobarbituric acid reactive substances (TBARS) and protein carbonyl content (PCC) were determined. Among all measured parameters spleen tissue exhibited a greatest difference between capons and intact pheasants. It was found the significantly higher concentration of PCC in capon’s spleen, but also a higher TSOD activity and strong tendency to higher GSH-Px, Mn- SOD and CuZn-SOD activity. Contrary, in the capon’s hart tissue TBARS concentration was significantly lower. The results indicated that pheasant capons maintained their antioxidant defence in almost all investigated tissue. Documented rise in spleen PCC content of capons might indicate on body defense activity with naturally produced free radicals utilized by the immune system.

Kako se sigurnosti hrane i zaštiti zdravlja potrošača pridaje sve veća pozornost, a prema dosadašnjim spoznajama zdravlje životinja preduvjet je kvalitetne i sigurne hrane, cilj ovoga istraživanja je sustavnim praćenjem i podizanjem zdravlja životinja postići bolja reprodukcijska i proizvodna svojstva te povećati sigurnost hrane. Budući je intenzivna proizvodnja često povezana s različitim stresorima (hranidbeni, okolišni, oksidacijski itd.), istražen je učinak korekcije obroka i primjene različitih dodataka u hranu za životinje (antioksidansa i prirodnih imunomodulatora) na poboljšanje zdravlja i prinosa životinja (perad, preživači, svinje) te boljeg nutricionističkog sastava, probavljivosti i dijetetske vrijednosti gotovog proizvoda. Rezultati istraživanja provedenih na različitim vrstama i pasminama životinja primijenit će se u poboljšanju i standardizaciji proizvodnje s ciljem izrade specifikacije neophodne za dobivanje oznake zemljopisnoga podrijetla ili oznake izvornosti.

Mlijeko kao animalni proizvod iznimno je važno u prehrani ljudi. S druge strane, veliki udio mliječne masti čine zasićene masne kiseline, čija je konzumacija povezana s pojavom bolesti srčanog i krvožilnog sustava. U našem smo istraživanju hranidbom krava lanenim sjemenom bogatim višestruko nezasićenom α-linolenskom masnom kiselinom pokušali modificirati sastav mliječne masti i na taj način učiniti mlijeko još prihvatljivijim za ljudsku upotrebu

Despite considerable progress, a routine clinical hematology is still based on identification of morphological quantitative and qualitative changes of peripheral blood cells and hematopoietic organs. A basic instrument is the stained blood smear and the accuracy of obtaining information requires the correct technique of making smears in order to avoid possible confusing artifacts and disarranged distribution of cells which leads to serious mistakes of differential counting and interpretation of findings. However, in veterinary medicine, the possibility of a mistake begins already during blood sampling. Animals not used to handling should be immobilized, by physical restraint or by using tranquillizers and anesthetics, which can cause changes in the number and appearance of red blood cells and platelets. Furthermore, although EDTA is the preferred anticoagulant for blood morphology studies for most animal species, EDTA causes hemolysis in blood samples obtained from some groups of birds and reptiles. In these animal species heparin is the most commonly used anticoagulant that interferes with proper staining of the blood cells. Evaluation of blood cell morphology in veterinary medicine requires a very long training and experience of veterinary practicians, since today not only domestic but also wild and exotic animals appear as patients. A wide range of specific morphologic characteristics of blood cells in different animal species are expressed. Starting with the red blood cells that are in most mammalian species in a shape of biconcave disc, but in camelids they are elliptic. Goats often have a spindle and fusiform erythrocytes, and in cervids a high percentage of erythrocytes are found in a shape of drepanocytes. Furthermore, erythrocytes of birds, fish and reptiles contain a nucleus. The dynamics of erythrocytes maturation also differ, as well as their release in the peripheral blood, so consequently, depending on animal species, different characteristics of regenerative blood film in a response to anemia are recognized. Regarding the leukocytes, the greatest morphologic diversity due to animal species and breeds we find in the eosinophils. By the number, appearance and intensity of staining eosinophilic granules, we can with great certainty know from which animal does the blood smear originate. A great challenge is to distinguish eosinophils from heterophils in small rodents, birds and reptiles. Heterophils in their role of defending the organism respond to neutrophils, but their granules are stained red. Basophils in birds and reptiles have a round nucleus, unlike the typical segmented nucleus in the mammalian. Morphologic characteristics of lymphocytes are very similar in all animals, while some shapes of monocytes nuclei are more common in particular animal species. Platelets in birds and reptiles have a nucleus and can be easily mistaken for lymphocytes by insufficiently experienced hematologists. In reptiles, another cell type is distinguished, the azurophiles, whose role and origin has not yet been fully understood. Therefore, due to the number and variety of animal species, in veterinary hematologic cytology, an extensive knowledge and experience is needed in identifying physiological blood cell morphology and it is a prerequisite for proper evaluation of morphological changes in various pathological conditions.

Computerized image analysis allows the numerical objectification of most suptile, often not promptly noticeable changes in cells. By using digital morphometry, changes in cells reduce the impact of the objective assessment on the cytological diagnosis and thus enhance the reproducibility and objectivity of the differentiation. Using digital images it allows measurement of diverse parameters in many structures. Therefore, the analysis is less expensive and it is carried out faster and more precise. In human cytology and histology morphometric parameters are used in comparison of normal cells in different physiological conditions as well as for reliable differentiation between malignant and benign cells of different tumors and also in comparison cells of primary and metastatic tumors. In veterinary medicine, blood cell characteristics between different animal species and in different physiological conditions were analyzed. Although erythrocytes of different bird species have the identical characteristics of cells with a nucleus, they differ significantly in size and shape within each species which is easily to distinguish by objectivising morphometric parameters (area, convex area, outline, maximum and minimum radius, length and breadth, as well as form factor and elongation factor). Our results showed that large birds, such as ostriches, have the largest cells with relatively small nuclei in comparison to other species which can be seen in smaller nucleo-cytoplasmic proportion, their nuclei are more extended and more unsymmetrical. Turkey erythrocytes are second in area, outline and convexity but with smaller nuclei area that are more extended. Hen erythrocytes are the smallest but with relatively largest nuclei in comparison to the cytoplasm as well as they are the most rounded and most symmetrical. Duck erythrocyte characteristics are somewhere between the other three more clearly defined species. The differences were also recognized regarding the gender in New Zealand white rabbits. The results showed that erythrocytes of females had a larger area, greater maximal and minimal radius, greater convex area and greater length and cell elongation factor than the erythrocytes of males. Very interesting results were obtained by morphometric analyze of lymphatic cells in the peripheral blood during different methods of vaccination. The results showed that the peripheral blood lymphocytes of chickens vaccinated by means of nebulisation compared to those parenteraly vaccinated were significantly larger in most examinated variables: area, outline, minimal and maximal radius, convex area, length and breadth and form-factor as well as the elongation factor of the cell as a whole (regarding lymphocyte cytoplasm) and nuclei, after vaccination, indicating the significantly higher metabolic activity of cells. In the lymphocytes of parenteraly vaccinated chickens, only the nucleo- cytoplasmic ratio was higher, indicating lower metabolic activity. Morphometric analysis of normal erythrocytes can be the basis for elaboration of reference values for each animal species. Such numerical reference values would provide more quality standard for determination of aberrations in physiological and pathological conditions as well as more precise starting premise in diagnostics of hematological disturbances and diseases in birds. Furthermore, morphometric characteristics of immunocompetent cell transformation can be used to evaluate immune responses to vaccination and vaccine efficacy.

The Effects of Splenectomy and Autologous Spleen Transplantation on Complete Blood Count and Cell Morphology in a Porcine Model

Marek's disease is a common lymphoproliferative disease of chickens, usually characterized by mononuclear cell infiltration of the peripheral nerves and various other organs and tissues including the iris and skin. The group of 70 newly-hatched chicks was vaccinated by means of nebulisation and exposed for 60 seconds to the HVT FC 126 vaccine, while the other group of 70 chicks received the same vaccine by parenteral route (s.c.). The aim of this study was to compare the morphological patterns of the chicken's lymphocytes in their peripheral blood, before and after vaccination, either by means of nebulisation or parenteral injection. Image analysis (area, convex area, outline, maximum and minimum radius, length and breadth, as well as form factor and elongation factor) was performed using Sform software (VAMSTEC, Zagreb, Croatia). A total of 50 blood smears of vaccinated chickens were analysed (20 by means of nebulization and 30 by parenteral injection), with an average of 100 cells per smear. Statistical data analysis was performed by use of the Statistica 7.1. The results showed that the peripheral blood lymphocytes of chickens vaccinated by means of nebulisation comparedto those parenteraly vaccinated were significantly larger in most examinated variables: area, outline, minimal and maximal radius, convex area, length and breadth and form-factor as well as the elongation factor of the cell as a whole (regarding lymphocyte cytoplasm) and nuclei, after vaccination, indicating the significantly higher metabolic activity of cells. In the lymphocytes of parenteraly vaccinated chickens, only the nucleo-cytoplasmic ratio was higher, indicating lower metabolic activity. The results show that the morphological patterns of the immunocompetent cell transformation could be used to evaluate immune responses to vaccination and vaccine efficacy. We conclude that nebulisation as a mode of vaccination against Marek’s disease stimulates the transformation of immunocompetent cells much earlier, thus shortening the time of immunosupresion and improving immune response. This is of paramount importance for the practical use of Marek’s disease vaccine.

Marekova bolest predstavlja jedan od najvećih problema u proizvodnji peradi pri čemu je cijepljenje jedan od najvažnijih postupaka njena suzbijanja. U istraživanju se rabilo cjepivo od soja FC 126 herpesvirusa purana primijenjeno jednodnevnim pilićima postupkom nebulizacije, te su se analizirale morfometrijske značajke limfocita u perifernoj krvi pilića prije cijepljenja te 3., 4., 7., 10., 14. i 21. dan nakon cijepljenja. Statistički značajne razlike nađene su u svim istraživanim pokazateljima: površini, opsegu, minimalnom i maksimalnom polumjeru, ispupčenosti odnosno konveksnosti površine, duljini i širini te pravilnosti i izduženosti cijele stanice, odnosno citoplazme limfocita kao i u nukleo-citoplazmatskom omjeru. Istovjetno su istraženi i parametri jezgre: površinu, opseg, minimalni i maksimalni polumjer, pravilnost i izduženost. Veličina limfocita se trećeg dana nakon cijepljenja značajno smanjila u odnosu na njihovu veličinu prije cijepljenja, dok su četvrtog dana limfociti bili najmanji. Ujedno su se smanjili i svi ostali mjereni pokazatelji: površina i opseg, duljina i širina, minimalni i maksimalni polumjer te konveksnost cijelih stanica i jezgara. Nakon toga su limfociti postupno rasli, te bili najveći 21. dana nakon cijepljenja. Istodobno se povećavala površina, opseg, minimalni i maksimalni polumjer citoplazme i jezgara. Pravilnost stanica se tijekom ispitivanog razdoblja također mijenjala, te su limfociti bili najpravilnijeg oblika 3., 7. i 21. dana. Jezgre su bile pravilnije, podjednako od 4. do 14. dana, a najpravilnije 21. dana. Tijekom istraživanog razdoblja omjer jezgre i citoplazme bio je podjednak sve do 14. dana kada je ustanovljen njegov značajan porast, te se potom, 21. dana vratio na ustaljene vrijednosti. Postignuti rezultati pokazuju morfološke značajke transformacije imunokompetentnih stanica, koje se mogu rabiti u procjeni imunog odziva na cijepivo, učinka cjepljenja i mogućeg docjepljivanja.

This study evaluates the effects of total splenectomy and autologous spleen transplantation in a porcine model on complete red blood cell count, platelet count, percentage of reticulocytes and red blood cell morphology. Nineteen pigs observed in the research were randomly divided into three groups. The first group underwent sham surgery, the second group was splenectomized and the third group, after being splenectomized, had 20 % of autologous spleen tissue transplanted into the greater omentum. Blood was sampled prior to surgery and on the 1st, 5th, 12th and 26th days after surgery. Erythrocyte count, indices and morphology, hemoglobin, hematocrit and platelet count were assayed. Erythrocyte count was significantly decreased postoperatively in sham-operated pigs (day 5) and in autotransplanted pigs (days 5 and 12), while in splenectomized pigs a significant decrease of erythrocyte count was present on all postoperative days of the experiment. Hemoglobin values and hematocrit changed according to changes in red blood cell count during the whole period of the experiment. Erythrocyte indices of splenectomized pigs were significantly increased postoperatively (MCV on the 12th and 26th days, MCH and MCHC on day 5). Autotransplanted pigs had significantly lower MCV (days 1, 5 and 12) and MCH (days 1 and 12), but significantly higher MCHC (5th day). The platelet count was significantly increased (12th day) in splenectomized pigs. The percentage of reticulocytes in the blood, including polychromasia, was significantly increased in all three groups of pigs on all postoperative days of the experiment. Morphological abnormalities of erythrocytes were found on the blood smears of all investigated groups of pigs, with prevalence on the smears of splenectomized pigs. The anemia and retikulocytosis found in blood samples of all investigated groups of pigs may have been physiological results showing the classical postoperative organism reaction to blood loss and surgical trauma. The more frequent appearance of variations in red blood cell morphology, such as the appearance of leptocytes, codocytes and Howell-Jolly bodies in splenectomized pigs when compared with the other two experimental groups, suggests that this was a change specific for splenectomy.

Marek's disease is one of the major problems in poultry production today and specific vaccination is one of the most important modes of its prevention. The aim of this study was to compare morphological patterns of chicken's lymphocytes in peripheral blood, before and on the 3rd, 4th, 7th, 10th, 14th and 21st day after vaccination by means of either nebulization or parenteral injection. Image analysis (area, convex area, outline, maximum and minimum radius, length and breadth, as well as form factor and elongation factor) was performed using the SFORM software (VAMSTEC, Zagreb). A total of 50 blood smears of vaccinated chickens (20 by means of nebulization and 30 by parenteral injection), with an average of 100 cells per smear, were analysed. The results showed that peripheral blood lymphocytes of chickens vaccinated by means of nebulization compared with parenteraly vaccinated were significantly larger in most examinated variables: area, outline, minimal and maximal radius, convex area, length and breadth, form factor as well as elongation factor of the whole cell (regarding lymphocyte cytoplasm) and nuclei, on the 3rd, 4th, 7th, 10th, 14th and 21st day after vaccination. In lymphocytes of parenteraly vaccinated chickens, only the nucleo-cytoplasmic ratio was higher. The results show morphological patterns of immunocompetent cell transformation, which can be used to evaluate immune responses to vaccination and vaccine efficacy. We conclude that nebulization as a mode of vaccination against Marek’s disease stimulates the transformation of immunocompetent cells much earlier, thus shortening the time of immunosupresion and increasing the immune response.