Development of a Transgenic Flammulina velutipes Oral Vaccine for Hepatitis B

Oralna primjena cjepiva dobivenih iz gljiva obećavajuća je za prevenciju zaraznih bolesti. Jestive gljive smatraju se prikladnim domaćinom za proizvodnju cjepiva zbog malih troškova proizvodnje i malog rizika kontaminacije gena. Međutim, zbog slabe ekspresije antitijela gljive imaju limitirani potencijal za razvoj oralnih cjepiva. Niska razina ekspresije gena vjerojatno je posljedica nečistoće transgenih micelija, budući da se za transformaciju uglavnom koriste miceliji dikariona. U radu je transformacijom gena iz gljive Flammulina velutipes pomoću bakterije Agrobacterium dobiven stabilni antigen hepatitisa B (HBsAg). Zatim je uslijedilo formiranje plodišta i klijanje bazidiospora. Metodom Western blot potvrđeno je formiranje HBsAg. Razine ekspresije HBsAg u plodištima gljive F. velutipes bile su: (129,3±15,1), (1 10,9±1,7) i (161,1±8,5) ng po gramu ukupnog topljivog proteina. Međutim, te vrijednosti mogu biti i veće, jer u radu nije postignuta potpuna ekstrakcija proteina. Dvije od četiri svinje u pokusnoj skupini imale su IgG protutijela na HBsAg nakon hranidbe plodištima transgene gljive F. velutipes tijekom 20 tjedana. U uzorcima krvi svinja u kontrolnoj skupini nisu opažena protutijela na HBsAg. Jedna od dviju pozitivnih svinja imala je vrijednosti titra protutijela na HBsAg od 5,36 miliinternacionalnih jedinica po mililitri (mlJ/mL) u desetom tjednu i 14,9 mlJ/mL u četrnaestom tjednu ispitivanja, nakon čega su se te vrijednosti smanjivale. Druga je svinja imala vrijednosti titra od 9,75 mlJ/mL u četrnaestom tjednu, 17,86 mlJ/mL u osamnaestom tjednu i 39,87 mlJ/mL u dvadesetom tjednu ispitivanja. Imunogenost svinja hranjenih plodištima transgene gljive F. velutipes potvrđuje mogućnost primjene ove gljive kao oralnog cjepiva.Orally administered fungal vaccines show promise for the prevention of infectious diseases. Edible mushrooms are deemed appropriate hosts to produce oral vaccines due to their low production cost and low risk of gene contamination. However, their low expression level of antigens has limited the potential development of oral vaccines using mushrooms. The low expression level might result from impurity of the transgenic mycelia since dikaryotic mycelia are commonly used as transformation materials. In this study, stable transgenic hepatitis B virus surface antigen (HBsAg) in Flammulina velutipes transformants was obtained by Agrobacterium-mediated transformation, followed by fruiting and basidiospore mating. The formation of HBsAg was detected by western blot analysis. The expression levels of HBsAg in transgenic F. velutipes fruiting bodies were (129.3±15.1), (110.9±1.7) and (161.1±8.5) ng/g total soluble protein. However, the values may be underestimated due to incomplete protein extraction. Two of the four pigs in the experimental group produced positive anti-HBsAg-specific IgG after being fed the HBsAg transgenic F. velutipes fruiting bodies for 20 weeks, while no anti-HBsAg antibody was detected in the control group. One of the positive pigs had HBsAg titres of 5.36 and 14.9 mIU/mL in weeks 10 and 14, respectively, but expression faded thereafter. The other positive pig displayed HBsAg titres of 9.75, 17.86 and 39.87 mIU/mL in weeks 14, 18 and 20, respectively. The successful immunogenicity in pigs fed transgenic F. velutipes fruiting bodies demonstrated the potential of using the fungus as an oral vaccine

Excavatoids O and P, New 12-Hydroxybriaranes from the Octocoral Briareum excavatum

Two new 12-hydroxybriarane diterpenoids, designated as excavatoids O (1) and P (2), were isolated from the octocoral Briareum excavatum. The structures of briaranes 1 and 2 were established on the basis of extensive spectral data analysis. Excavatoid P (2) is the first metabolite which possesses a 6β -chlorine atom in briarane analogues

Electromagnetic Wave Theory and Applications

Contains table of contents for Section 3, reports on ten research projects and a list of publications.National Aeronautics and Space Administration Contract 958461U.S. Navy - Office of Naval Research Grant N00014-92-J-1616U.S. Navy - Office of Naval Research Grant N00014-89-J-1019U.S. Navy - Office of Naval Research Grant N00014-90-J-1002U.S. Army Cold Regions Research and Engineering Laboratory Contract PACA89-95-K-0014Mitsubishi Corporation Agreement Dated 8/31/95U.S. Navy - Office of Naval Research Grant N00014-92-J-4098U.S. Federal Aviation Administration Grant 94-G-007DEMACO Corporation Contract DEM-95-MIT-55Joint Services Electronics Program Contract DAAH04-95-1-003

Electromagnetic Wave Theory and Applications

Contains table of contents for Section 3, reports on three research projects and a list of publications.California Institute of Technology/Jet Propulsion Laboratory Contract 959548National Aeronautics and Space Administration Grant NAGW-1617National Aeronautics and Space Administration Grant Contract 958461U.S. Navy - Office of Naval Research Grant N00014-92-J-1616U.S. Navy - Office of Naval Research Grant N00014-92-J-4098Digital Equipment Corporation AGMT DTD 11/16/93Joint Services Electronics Program Contract DAAL03-92-C-0001Joint Services Electronics Program Grant DAAH04-95-1-0038MIT Lincoln Laboratory P.O. No. BX-5424U.S. Navy - Office of Naval Research Grant N00014-90-J-1002U.S. Navy - Office of Naval Research Grant N00014-89-J-1019DEMACO Agreement 11/15/93Federal Aviation Administration Grant 94-G-007U.S. Army Cold Regions Research and Engineering Laboratory Contract DACA89-93-K-000

HBsAg Profiles in Patients Receiving Peginterferon Alfa-2a plus Ribavirin for the Treatment of Dual Chronic Infection with Hepatitis B and C Viruses

Background. With use of peginterferon alfa-2a and ribavirin combination therapy in patients with dual chronic hepatitis B virus (HBV) and hepatitis C virus (HCV) infection, 11.2% of patients achieved clearance of hepatitis B surface antigen (HBsAg) at 6 months after treatment; however, reactivation of HBV DNA was observed in 36.3%. We investigated the predictive potential of HBsAg quantification. Methods. HBsAg quantification was performed in 120 e antigen-negative patients dually infected with HBV and hepatitis C virus and treated with peginterferon alfa-2a/ribavirin for 48 weeks (HCV genotype 1; ) or n p 74 24 weeks (HCV genotype 2/3; ). HBsAg was quantified at baseline, week 4, week 12, end of treatment, and n p 46 24 weeks after treatment. Results. The baseline median serum HBsAg level was 120 IU/mL and decreased gradually during treatment. Low baseline HBsAg was significantly associated with HBsAg clearance (40% for HBsAg level р20 IU/mL vs 2.2% for HBsAg level 120 IU/mL; ). A decrease in HBsAg level from baseline to week 12 of 50% was associated P ! .05 with a reduced likelihood of HBV DNA reactivation in patients with baseline undetectable serum HBV DNA (positive predictive value, 89.5%). Conclusions. HBsAg quantification appears to be a useful indicator of posttreatment outcome in patients dually infected with HBV and hepatitis C virus