Isolation of highly pathogenic avian influenza H5N1 from table eggs after vaccinal break in commercial layer flock.

In May 2009, during routine monitoring of a commercial layer flock of about 87,000 birds kept in cages in 4 different houses that had been vaccinated 3 times with an inactivated H5N1 vaccine at weeks 1, 7, and 16, highly pathogenic avian influenza (HPAI) virus of subtype H5N1 was isolated and detected by real-time reverse transcriptase-polymerase chain reaction (RT-PCR) in tracheal and cloacal swabs collected from houses 3 and 4; 7 days after onset of clinical signs, there was an increase in mortality accompanied by a decrease in egg production and egg quality. In addition, using RT-PCR, the viral RNA could be detected from albumin and eggshell as well. Seven days after the onset of the clinical signs, the hemagglutination inhibition (HI) titers in the affected houses were 3.2 and 1.9 log2. In the other two houses, there were no clinical signs, and all tested samples were negative using virus isolation and real-time RT-PCR. The HI titers were 6.6 and 7.0 log2 in nonaffected houses. The isolated virus from egg albumin showed high nucleotides and amino-acid identities and clustered with viruses from recently H5N1-confirmed human infections and poultry from different places in Egypt. Moreover, several amino-acid substitutions of viral H5 protein were observed. The vaccinal break seems to be associated with immune escape mutants and/or improper vaccination. The role of contaminated eggs as a source of infection and as a vehicle for spread of the virus should be considered in area with avian influenza outbreaks.

Identification and characterization of a 26- to 28-kDa circulating antigen of Fasciola gigantica.

As a disease of domestic ruminants, fascioliasis is of considerable economic importance. Although serological tests are available for the diagnosis of the disease, they are of generally low specificity because of cross-reactivity with antigens from other parasites. There is a need to identify other Fasciola antigens on which more specific tests could be based. In the present study, a specific rabbit anti-serum and western-blot analyses were used to demonstrate the presence of a highly reactive antigen of 26-28 kDa not only in an extract of adult F. gigantica but also in the excretory/secretory products of the worms and in the bile secretions and sera of cattle that were naturally infected with this parasite. The 26- to 28-kDa antigen was isolated from preparative polyacrylamide gels, by electro-elution. The purified antigen showed a single peak at 5.8 min when analysed by capillary zone electrophoresis. It was characterized as protein containing 47.5% hydrophilic and 29.3% hydrophobic amino acids. Immunostaining demonstrated that the target epitope was located in the gut and tegument of adult F. gigantica and within the bile ducts, the portal tracts of the livers and the mucosa and muscularis of the gallbladders of infected cattle. A simple and rapid dot-ELISA technique based on the specific rabbit anti-serum was 100% specific when tested on the sera from nine cattle infected with F. gigantea and 27 uninfected cattle. In conclusion, the 26- to 28-kDa Fasciola antigen may be a promising candidate for the immunodiagnosis of fascioliasis.