Trajanje imunosti nakon cijepljenja s Avishield IB H120, cjepivom protiv ptičjeg zaraznog bronhitisa

The most effective means of controlling infectious bronchitis in poultry is by vaccination. Live attenuated vaccines based on the H120 strain (Mass serotype) are the most commonly used vaccines. Since vaccination is often performed in the hatchery, long duration of immunity is preferred. Avishield IB H120, a live attenuated vaccine based on the strain H120 is registered across the European Union and other countries, and has a declared immunity period of up to 5 weeks after vaccination. This study presents the results of an additional survey demonstrating protection against challenge with the IBV strain M41 up to 8 weeks after a single vaccination with Avishield IB H120. Seven weeks after vaccination, 94% of chickens vaccinated by spray were protected against the challenge. Eight weeks after vaccination, 80% of spray vaccinated chickens and 75% of orally vaccinated chickens were still protected against the challenge with virulent IBV.Cijepljenje je najučinkovitiji način kontrole zaraznog bronhitisa u peradi. Najčešće korištena cjepiva su živa atenuirana cjepiva temeljena na soju H120 (pripadaju Mass serotipu). Budući da se cijepljenje često provodi već u valionici, poželjno je dugo trajanje imunosti. Avishield IB H120, živo atenuirano cjepivo koje sadrži cjepni soj H120, registrirano je u brojnim zemljama Europske unije i na drugim tržištima s deklariranim trajanjem imunosti od 5 tjedana nakon cijepljenja. U ovom radu prikazani su rezultati dodatne studije gdje smo dokazali zaštitu od izazivačke infekcije s patogenim virusom M41 i 8 tjedana nakon cijepljenja s Avishield IB H120 putem spreja ili vode za piće

Identification of Small Molecule Inhibitors of Pseudomonas aeruginosa Exoenzyme S Using a Yeast Phenotypic Screen

Pseudomonas aeruginosa is an opportunistic human pathogen that is a key factor in the mortality of cystic fibrosis patients, and infection represents an increased threat for human health worldwide. Because resistance of Pseudomonas aeruginosa to antibiotics is increasing, new inhibitors of pharmacologically validated targets of this bacterium are needed. Here we demonstrate that a cell-based yeast phenotypic assay, combined with a large-scale inhibitor screen, identified small molecule inhibitors that can suppress the toxicity caused by heterologous expression of selected Pseudomonas aeruginosa ORFs. We identified the first small molecule inhibitor of Exoenzyme S (ExoS), a toxin involved in Type III secretion. We show that this inhibitor, exosin, modulates ExoS ADP-ribosyltransferase activity in vitro, suggesting the inhibition is direct. Moreover, exosin and two of its analogues display a significant protective effect against Pseudomonas infection in vivo. Furthermore, because the assay was performed in yeast, we were able to demonstrate that several yeast homologues of the known human ExoS targets are likely ADP-ribosylated by the toxin. For example, using an in vitro enzymatic assay, we demonstrate that yeast Ras2p is directly modified by ExoS. Lastly, by surveying a collection of yeast deletion mutants, we identified Bmh1p, a yeast homologue of the human FAS, as an ExoS cofactor, revealing that portions of the bacterial toxin mode of action are conserved from yeast to human. Taken together, our integrated cell-based, chemical-genetic approach demonstrates that such screens can augment traditional drug screening approaches and facilitate the discovery of new compounds against a broad range of human pathogens

Genome-wide identification of Pseudomonas aeruginosa virulence activities and their small molecule inhibitors using phenotypic screens in yeast

Bakterija Pseudomonas aeruginosa je oportunistički patogen u ljudi, visoko rezistentna na mnoge antimikrobne agense i može razviti visok stupanj višestruke rezistencije na antibiotike, zbog čega se javila potreba za otkrivanjem novih metoda pronalaska ciljnih molekula za djelovanje lijekova. U ovom radu je razvijen novi pristup korištenja kvaščevih stanica kako bi se identificirali i okarakterizirali potencijalni ciljni geni bakterije P. aeruginosa PAO-1. Koristeći GatewayTM tehnologiju, zbirka 505 PAO-1 gena, uključenih u esencijalne stanične procese i virulenciju, bila je prebačena u kvaščev vektor koji je sadržavao inducibilan promotor. Nakon transformacije kvasca Saccharomyces cerevisiae, kvaščeve stanice su bile nacijepljene na podlogu na kojoj je bila inducirana ekspresija PAO-1 gena. Koristeći ovaj pristup, selektirana su četiri bakterijska toksina i šest esencijalnih gena kao potencijalne ciljne molekule za djelovanje lijekova. Geni koji su uzrokovali inhibiciju rasta kvaščevih stanica se trenutno koriste u drugom pretraživanju, kako bi se identificirali kemijski spojevi koji mogu revertirati inhibiciju rasta. Ovaj pristup, temeljen na korištenju cijelih kvaščevih stanica će proširiti primjenu "high through-put" tehnika funkcionalne genomike na patogene mikroorganizme kojima se teško eksperimentalno upravlja.Pseudomonas aeruginosa is an opportunistic human pathogen, highly resistant to multiple antimicrobial agents and can develop high-level multidrug resistance, highlighting the crucial need for new drug target discovery methods. A novel yeast-based approach was developed to identify and characterize potential drug target genes of P. aeruginosa PAO-1. Using the GatewayTM technology, a collection of 505 selected PAO-1 ORFs involved in essential cellular processes and virulence, was transfered to a yeast vector containing an inducible promoter. After transformation in S. cerevisiae, yeast colonies were re-arrayed onto plates where the expression of PAO-1 ORFs was induced. Using such whole-cell approach, four bacterial toxins and six essential genes were selected as potential drug targets. These genes causing growth inhibition are currently being used in a second screen designed to identify chemical compounds capable of reverting the growth inhibition. Such a yeast-based approach will extend the application of high-throughput functional genomics techniques to pathogenic microorganisms that are not readily manipulated experimentally

Rapid transient expression of human granulocyte-macrophage colony-stimulating factor in two industrial cultivars of tobacco (Nicotiana tabacum L.) by agroinfiltration

We report the production of hGM-CSF cytokine in leaves of industrial tobacco cultivars DH-17 and DH-27 by using Agrobacterium-mediated transient expression. We prove the concept that very high biomass industrial tobacco plants are suitable platforms for rapid, low cost production of foreign proteins. Successful transient expression of the GM-CSF was achieved in less than three months, opening the possibility for future applications of this approach in rapid response production of various proteins of non-plant origin in industrial tobacco

Safety and efficacy profile of live, intermediate plus vaccine against infectious bursal disease based on strain G6

Abstract Infectious bursal disease (IBD) is an acute, highly contagious, immunosuppressive disease of young chickens that causes considerable economic loss in the poultry industry worldwide. Vaccination with live attenuated vaccines is still the most important method used for the control and prevention of IBD in chickens. Here we present the results of in vitro characterization, as well as efficacy and safety testing of a live, intermediate plus vaccine against IBD based on strain G6. Strain characterization confirmed that G6 strain is an intermediate plus strain, showing a high degree of homology with the existing vaccine strains of the same virulence. Safety studies showed that chickens can be vaccinated from 10 days of age. Onset and duration of immunity in specific pathogen free and maternally derived antibodies (MDA) chickens was proven to be 14 and 35 days after vaccination, respectively. When immunizing MDA-positive chickens, vaccine is capable of breakthrough at a titer of ≤500 ELISA units. The field trial conducted on commercial broilers showed a 95% protection against vvIBDV challenge. Stability of the freeze-dried vaccine after reconstitution was confirmed over a period of 3 h. Overall, IBD G6 vaccine has shown good safety and efficacy profile in accordance with European Pharmacopoeia requirements