Isolation and molecular characterization of novel IBV isolates from broiler chicken farms in Egypt

A survey of infectious bronchitis virus [IBV] genotypes in 25 commercial broiler flocks of various ages [20-35 days] raised in Al-Behera and Kafr-Elsheikh governorates and suffering from respiratory symptoms and pathological changes in kidney associated with high mortality rate. All flocks were vaccinated against IB disease. Tissue samples [trachea, lung and kidney] were collected aseptically from these flocks. Then virus propagation was performed in embryonated SPF chicken eggs via allantoic sac inoculation. Results of virus isolation trails from the collected organs revealed 15 IBV isolates [60%] out of 25 flocks as judged by antigen detection in CAMs by the AGPT against reference IBV Beaudette antiserum after 2-5 chicken embryo serial passages. However, three of 15 IBV isolates were also positive by the slide HA test. Moreover, 5 flocks gave positive slide HA test and negative by the AGPT. The other 5 flocks gave negative slide HA test and AGPT. Then selected ten IBV field isolated strains in this study were characterized by RT-PCR [all of ten selected isolates are positive for S1 gene], and then sequence analysis of partial S1 spike glycoprotein gene of seven IBV field isolates in this study [11, 15, 21, 13, 19, 22, 24] were made. The seven IBV field isolates showed 97% to 98.3 % and 96.7% to 98.3% nucleotide sequence identity to IBV-CU-2-SP1 and Eg/12120s/2012 strain [variant 2 like strain], respectively. Nucleotide identity between these seven IBV isolates ranged from 97.7% to 99% and between these isolates and vaccinal strain used in Egypt [M41, H120, Ma5, 4/91, CR88 and D274] ranged from 64.7% to 65.7%, 65.3% to 66.3%, 65.7% to 66.7%, 67.3% to 68.3%, 68.6% to 69.6% and 84.2% to 84.8%, respectively. The presence of these strains may account for vaccination failure against IBV, since all IBV isolates were from vaccinated chickens. This study demonstrates a constant evolution of IBV in Egypt that necessitates continuous monitoring to control the spread of infections, and the development and use of vaccines based on indigenous viruses

Epidemiological studies on highly pathogenic avian influenza H5N1 during late 2009 and 2010 in Egypt

Epidemiological studies on AI virus H5N1 in different governorates in Egypt [Alexandria, Bohera, Cairo, El-fayom, Gharbia, Giza and Dakahlia] during late 2009 and 2010 were carried out. These studies included seventy five flocks [49 broilers, 17 layer, 4 breeder and 5 duck flocks] showing high mortality rate with respiratory manifestations and diarrhea, through isolation in specific pathogen free eggs and identification by hemagglutination inhibition test using H5N1 antiserum. The data revealed the following: a percentage of 32.7%, 76.5%, 50% and 40% in broiler, layer, breeder and duck flocks respectively were positive for AIV H5N1. The highest incidence was recorded in layer flocks followed by broiler breeder flocks, duck then broiler flocks with total incidence of 44% in all species. Sequencing and phylogenetic tree of hemagglutinin [HA] gene of six positive AIV H5N1 isolates from chickens during 2010 were done. Phylogenetic tree showed that all HA gene sequences blonged to highly diverse clade 2.2.1 viruses according to WHO/FAO/OIE nomenclature. Analysis of amino acid sequences of HA glycoprotein revealed some mutations at the receptor binding site of the HA molecule

Factors affecting hemagglutination activity of avian influenza virus subtype H5N1

Hemagglutination [HA] and hemagglutination inhibition [HI] tests for avian influenza [AI] virus [H5N1] were standardized using varying various factors like erythrocytes from different species, type of diluent, incubation temperature and incubation period. The virus was peropagated in embryonated chicken eggs [9-11 days]. The allantoic fluid [AF] was harvested 36 hours post incubation and was confirmed by slide hemagglutination test. The maximum HA titres were obtained using 1% RBCs of chicken, sheep, duck, geese, pigeon, quails and turkey for 30-40 minutes. The haemagglutination activity showed best titer when used phosphate buffer saline than normal saline and also when incubated at 25[degree sign] C instead of 37[degree sign] C