Primjena kompetitivne ekskluzije u sprečavanju zaraze bakterijom Salmonella enterica subspecies enterica serovar Enteritidis u pilića.

A total of 250 newly-hatched broiler chickens allocated to two groups were used and two replicate trials were carried out. The chicks were dosed by gavage with commercial competitive exclusion (CE) product (trial groups) or with 0.5 ml of physiological saline (controls) at day 0, day 1 and day 5. On day 2, eight birds per tier were challenged by oral gavage with 0.5 ml of an approximate 107 organism/ml dilution of S. Enteritidis phage type 9a and were used as seeders. On the fifth day after challenge, and at 14 and 21 days of age, 12 chicks from each group were selected and Salmonellae were counted in their caeca. On days 14 and 21 the proportion of infected animals was significantly lower (P0.1) among treated groups with regard to the spleen colonization observed on days 14 and 21. On day 14, the number of chickens with infected spleen was 4/24 in challenge control and 9/24 in the treated group. On day 21 this proportion was 4/24 and 3/24, respectively. Commercial competitive exclusion product provided a significant protective effect against pathogenic S. Enteritidis PT 9a used for challenge.Ukupno 250 netom izleženih tovnih pilića korišteno je u dva usporedna istovjetna pokusa. Pilićima je u preporučenoj dozi primijenjen komercijalni proizvod za kompetitivnu ekskluziju (CE) u dobi 0, 1 i 5 dana (pokusna skupina) ili fiziološka otopina (kontrolna skupina). Drugog dana, 16 pilića bilo je zaraženo bakterijom Salmonella Enteritidis fagotipom 9a (suspenzijom s približno 107 bakterija/ml). Ti pilići bili su upotrijebljeni kao izvor zaraze i smješteni zajedno s ostalim pilićima pokusne odnosno kontrolne skupine. Petog dana nakon zaražavanja, tj. u dobi 14 i 21 dan, izdvojeno je 12 pilića po skupini kojima je određen broj salmonela u sadržaju cekuma. Udio zaraženih pilića bio je značajno manji (P<0,05) 14. i 21. dana u skupini koja je primila proizvod CE u odnosu na kontrolnu skupinu. Od ukupnog broja pilića pokusne skupine, u 50% dokazana je infekcija 14. dana, dok se 21. dana taj broj smanjio na četvrtinu. Udio zaraženih pilića u kontrolnoj skupini iznosio je 21 odnosno 17 u odnosu na 14. i 21. dan. Salmonela je bila izdvojena 14. dana iz slezene 4 pileta kontrolne te iz 9 pilića pokusne skupine. Taj udio 21. dana iznosio je 4/24 u odnosu na 3/24. Ovim istraživanjem dokazan je znatan zaštitni učinak kompetitivne ekskluzije u sprečavanju zaraze patogenim sojem bakterije Salmonella Enteritidis fagotip 9a

Primjena kompetitivne ekskluzije u sprečavanju zaraze bakterijom Salmonella enterica subspecies enterica serovar Enteritidis u pilića.

A total of 250 newly-hatched broiler chickens allocated to two groups were used and two replicate trials were carried out. The chicks were dosed by gavage with commercial competitive exclusion (CE) product (trial groups) or with 0.5 ml of physiological saline (controls) at day 0, day 1 and day 5. On day 2, eight birds per tier were challenged by oral gavage with 0.5 ml of an approximate 107 organism/ml dilution of S. Enteritidis phage type 9a and were used as seeders. On the fifth day after challenge, and at 14 and 21 days of age, 12 chicks from each group were selected and Salmonellae were counted in their caeca. On days 14 and 21 the proportion of infected animals was significantly lower (P0.1) among treated groups with regard to the spleen colonization observed on days 14 and 21. On day 14, the number of chickens with infected spleen was 4/24 in challenge control and 9/24 in the treated group. On day 21 this proportion was 4/24 and 3/24, respectively. Commercial competitive exclusion product provided a significant protective effect against pathogenic S. Enteritidis PT 9a used for challenge.Ukupno 250 netom izleženih tovnih pilića korišteno je u dva usporedna istovjetna pokusa. Pilićima je u preporučenoj dozi primijenjen komercijalni proizvod za kompetitivnu ekskluziju (CE) u dobi 0, 1 i 5 dana (pokusna skupina) ili fiziološka otopina (kontrolna skupina). Drugog dana, 16 pilića bilo je zaraženo bakterijom Salmonella Enteritidis fagotipom 9a (suspenzijom s približno 107 bakterija/ml). Ti pilići bili su upotrijebljeni kao izvor zaraze i smješteni zajedno s ostalim pilićima pokusne odnosno kontrolne skupine. Petog dana nakon zaražavanja, tj. u dobi 14 i 21 dan, izdvojeno je 12 pilića po skupini kojima je određen broj salmonela u sadržaju cekuma. Udio zaraženih pilića bio je značajno manji (P<0,05) 14. i 21. dana u skupini koja je primila proizvod CE u odnosu na kontrolnu skupinu. Od ukupnog broja pilića pokusne skupine, u 50% dokazana je infekcija 14. dana, dok se 21. dana taj broj smanjio na četvrtinu. Udio zaraženih pilića u kontrolnoj skupini iznosio je 21 odnosno 17 u odnosu na 14. i 21. dan. Salmonela je bila izdvojena 14. dana iz slezene 4 pileta kontrolne te iz 9 pilića pokusne skupine. Taj udio 21. dana iznosio je 4/24 u odnosu na 3/24. Ovim istraživanjem dokazan je znatan zaštitni učinak kompetitivne ekskluzije u sprečavanju zaraze patogenim sojem bakterije Salmonella Enteritidis fagotip 9a

Promjene broja krvnih stanica u jednodnevnih pilića cijepljenih protiv Marekove bolesti sojem FC 126 herpesvirusa purana postupkom nebulizacije.

Marek’s disease is one of the greatest problems in poultry production today and vaccination is one of the most important ways of prevention. An innvoative method of vaccine delivery by means of nebulisation was used for vaccination of newly hatched chicks using HVT FC 126 and its impact on blood cell count (WBC, RBC and differential cell count) and H/L ratio was measured. The trial was performed from day 1 to day 21 of life of newly hatched male chicks. Standard blood cell count was performed in a Neubauer hemocytometer, and differential cell count on blood smears stained with MayGrünwald-Giemsa. Blood for that purpose was taken from the jugular vein. The results show a significantly higher WBC count in the group vaccinated by means of nebulisation than in the non-vaccinated groups on day 5 of the trial, and from the group that received only physiological solution, on day 14. In the RBC count there were fluctuations but without any significant differences between the groups during the trial. In differential blood cell count, there were some significant differences on days 7 and 21 of the trial, but no differences in H/L ratio. These results show that vaccination by means of nebulisation significantly influenced the blood cell count but all changes were within the physiological range. Nebulisation as a method of vaccination could probably improve the immune response to the wild Marek’s disease virus, mimicking natural infection via the respiratory system and as a mass form of vaccination could be a powerful method for delivery of recombinant HVT vaccine.Marekova bolest predstavlja jedan od najvećih problema u uzgoju peradi pri čemu je cijepljenje jedan od najvažnijih postupaka suzbijanja. Korišten je novi postupak primjene cjepiva od soja FC 126 herpesvirusa purana jednodnevnim pilićima postupkom nebulizacije pri čemu je analiziran njegov utjecaj na broj stanica u krvi (broj leukocita, broj eritrocita i diferencijalnu krvnu sliku) kao i omjer neutrofila i limfocita (N/L). Pokus je proveden na jednodnevnim muškim pilićima od prvog do 21. dana života. Korišten je standardni postupak brojenja krvih stanica u Neubauerovoj komorici, dok je DKS načinjen na razmazima pune krvi obojene May-Grünwald-Giemsom. Krv za navedene postupke uzeta je iz jugularne vene. Rezultati pokazuju značajan porast broja leukocita u krvi kod skupine cijepljene postupkom nebulizacije u odnosu na necijepljene skupine petog dana pokusa, dok od skupine koja je primila samo fiziološku otopinu ima značajno više leukocita 14. dana pokusa. Broj eritrocita pokazuje varijacije u pokusnih skupina, ali bez značajnih razlika tijekom čitavog pokusa. Značajne razlike u diferencijalnoj krvnoj slici bile su ustanovljene 7. i 21. dana pokusa, dok kod N/L omjera nije bilo značajnih razlika. Navedeni rezultati pokazuju da je cijepljenje postupkom nebulizacije značajno utjecalo na broj krvnih stanica te moguće potaknulo specifičan imunosni odgovor, ali su sve promjene bile u fiziološkom rasponu. Nebulizacija kao postupak primjene cjepiva, iz navedenog, pretpostavlja se može poboljšati imunosni odgovor na virus Marekove bolesti oponašajući prirodan način zaražavanja putem dišnog sustava, a može biti izvrstan u masovnoj primjeni rekombinantnih cjepiva pripravljenih od herpesvirusa purana

Iskorjenjivanje enzootske leukoze goveda na farmi mliječnih krava primjenom različitih dijagnostičkih postupaka.

The aim of this study was to perform a diagnosis and molecular characterisation of bovine leukosis virus (BLV) on a Croatian dairy farm. For that purpose we compared the diagnostic tools used in our study, made sequence and phylogenetic analysis of our BLV and tried to describe the difficulties in the process of EBL eradication on the examined farm after reintegration into Croatian territory after the war. From 1998 to 2008 blood samples from a dairy farm in the North-eastern part of Croatia were tested serologically using AGID and ELISA. In 2002, 2003 and 2004 37%, 22% and 10% of animals were serologically positive, respectively. After the initial eradication steps, the disease reappeared in 2008, when all examined blood samples reacted positively in BLV-specific nested PCR. Finally, at the end of 2010, after an extended eradication program, which included the implementation of PCR together with regular ELISA testing for detection of positive animals, the farm obtained the status of “free of BLV”. Sequence and phylogenetic analysis of 524-nt fragment of BLV env gp51 encoding gene showed the greatest identity with the Croatian BLV genotype 8 isolates, so it was phylogenetically clustered with those isolates.Cilj istraživanja bio je dijagnosticirati i molekulski karakterizirati virus enzootske leukoze goveda (VELG) na jednoj hrvatskoj farmi mliječnih krava. U tu svrhu uspoređeni su dijagnostički postupci rabljeni u istraživanju te je napravljena analiza nukleotidnog slijeda VELG i filogenetska analiza. Opisane su i poteškoće u provođenju mjera iskorjenjivanja na pretraživanoj farmi tijekom mirne integracije ovog područja Hrvatske poslije domovinskog rata. Od 1998. do 2008. uzimani su uzorci krvi krava na farmi mliječnih krava smještenoj u sjeveroistočnom dijelu Hrvatske te serološki pretraživani gel difuzijskim precipitacijskim (GDP) i imunoenzimnim testom. U 2002. na VELG bilo je pozitivno 37%, 2003. 22%, a 2004. 10% životinja. Godine 2008. svi pretraženi uzorci krvi bili su pozitivni pretragom lančanom reakcijom polimerazom. Naposljetku, krajem 2010. nakon produženih mjera iskorjenjivanja što su uključivale primjenu lančane reakcije polimerazom istodobno s redovitim pretraživanjem imunoenzimnim testom kako bi se identificirale sve životinje pozitivne na ELG, farma je postala slobodna od ELG. Na osnovi filogenetske analiza odsječka VELG što kodira za gen env gp51 izolati su bili svrstani u istu skupinu (VELG genotip 8) s već istraženim hrvatskim izolatima VELG

Immunogenicity and Safety of La Sota Strain of Newcastle Disease Virus Administered to Newly Hatched Chicks by Nebulization

The objective of four trials performed on specific-pathogen-free and commercial chickens, either of light or heavy hybrids, was to evaluate the new vaccine delivery method to newly hatched chickens using commercial La Sota vaccine. The vaccine was given by means of nebulization using an ultrasonic device producing homologous aerosol of particles ranging 3–5 microns in diameter. Chickens were exposed to the La Sota vaccine for 30, 60 or 300 s in a closed chamber of the device, thus enabling constant particle size during vaccination. No adverse reaction to the given vaccine was recorded, and the immunity, developed no later than 7 days after vaccination, lasted for at least 49 days which was confirmed by challenge infection using Herts 33 strain of Newcastle disease virus. Maternal antibodies did not influence the development of immunity. Regarding the mode of vaccination, the described method is suitable for the control of Newcastle disease in both big poultry enterprises as well as small backyard flocks when newly hatched chickens are supplied from local hatcheries

Utjecaj cjepiva od soja FC 126 herpesvirusa purana primijenjenog postupkom nebulizacije i parenteralno na transformaciju imunokompetentnih stanica u pilića.

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.Marekova je bolest limfoproliferativna zarazna bolest kokoši, obično karakakterizirana mononuklearnim staničnim infiltacijama u perifernim živcima i ostalim organima i tkivima, uključujući šarenicu i kožu. Skupina od 70 netom izleglih pilića cijepljena je protiv Marekove bolesti cjepivom od soja FC 126 herpesvirusa purana postupkom nebulizacije izlaganjem aerosolu tijekom 60 sekundi, dok je druga skupina od 70 pilića primila isto cjepivo parenteralnim načinom (supkutano). Cilj ovog istraživanja bio je usporediti morfometrijske značajke limfocita u perifernoj krvi pilića prije i nakon cijepljenja kako postupkom nebulizacije tako i parenteralnim načinom primjene. Kompjutorska analiza slike učinjena je na osobnom računalu korištenjem programa „SFORM“ (Vamstec, Zagreb, Hrvatska). Ukupno je pretraženo 50 obojenih razmaza periferne krvi cijepljenih pilića (20 postupkom nebulizacije i 30 parenteralno), te je analizirano prosječno 100 limfocita po uzorku. Rezultati su pokazali da su limfociti u perifernoj krvi pilića cijepljenih postupkom nebulizacije u odnosu na parenteralno cijepljene bili značajno veći u većini istraživanih pokazatelja: površini, opsegu, minimalnom i maksimalnom polumjeru, dužini i širini, te ispupčenosti odnosno konveksnosti površine, u svim danima pokusa osim četvrtoga, što ukazuje na pojačanu metaboličku aktivnost tih stanica. U limfocitima parenteralno cijepljenih pilića samo je omjer jezgre i citoplazme bio veći. Postignuti rezultati pokazuju morfološke značajke transformacije imunokompetentnih stanica, koje se mogu rabiti u procjeni imunosnog odziva na cjepivo, učinka cijepljenja i mogućeg docjepljivanja. Zaključili smo da nebulizacija kao metoda cijepljenja protiv Marekove bolesti stimulira transformaciju imunokompetentnih stanica mnogo ranije, skraćujući time vrijeme imunosupresije i pojačavajući imunosni odziv. To je od osobite važnosti za praktičnu primjenu cjepiva protiv Marekove bolesti

Eradication of bovine leukosis virus on a diary farm through improved

virus detectio