Pathogenicity of Low Path H7N2 Avian Influenza Viruses from the Delmarva Peninsula for Broiler and Leghorn Chickens and Turkeys
B. S. Ladman, S. C. RosenbergerA, J. K. RosenbergerA, C. R. Pope, and J. Gelb, Jr.
Avian Biosciences Center, University of Delaware,
A AviServe LLC., 1 Innovation Way, Suite 1000,
SUMMARY. The vriulence of low pathogenicity (LP) type A H7N2 avian influenza virus (AIV) isolates recovered from chickens in Delaware and the eastern shore of Maryland in 2004 was evaluated. Three-week-old leghorn- and broiler-type chickens and turkeys were inoculated via the intraocular route with 103.5-104.0 50% embryo infectious dose (EID50) of virus per bird with, A/chicken/Delaware/Viva/04, A/chicken/Delaware/Hobo/04, and A/chicken/Maryland/Minh Ma/04. In broilers, the viruses produced respiratory signs, airsacculitis and microscopic lesions in the trachea and lung. In contrast, signs and lesions were less severe in turkeys and were rarely observed in specific-pathogen-free (SPF) leghorns. In broilers and SPF leghorns, AIV peaked on day 3 post inoculation (PI), based on virus isolation and real time reverse transcription polymerase chain reaction (RRT-PCR), and antigen capture testing. Infection in turkeys peaked on day 7 PI. Serum antibodies generally were detected earlier in broilers (day 7 PI) than in turkeys or SPF leghorns (day 14 PI) using agar gel immunoduffussion (AGID), hemagglutination-inhibition (HI) and the enzyme linked immunosorbent assay (ELISA).
A second trial was performed to further examine the disease susceptibility of the leghorn chicken given the comparatively mild responses noted in the first trial. A ten-fold higher dose of 104.5-105.0EID50 per chick given via the intraocular route was used. In addition, commercial-type leghorns were tested as were chicks from the SPF leghorn source. The higher AIV dose resulted in more rapid and consistent rates of infection and higher serum antibody responses in both types of leghorn chickens. However, as observed in the first trial, clinical signs and microscopic lesions in both types of leghorns were infrequent and very mild. These findings indicate leghorn-type chickens, which are commonly used for pathogenicity assessments because of their availability, may not be the most suitable host for evaluating the virulence potential of LP AIV.
J. Gelb Jr., B. S. Ladman, M. J. Licata, M. H. Shapiro, and L. R. Campion
Department of Animal and Food Sciences, Avian Biosciences Center,
SUMMARY. The potential for infectious bronchitis virus (IBV) and Newcastle disease virus (NDV) replication interference was evaluated using quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Fourteen-day-old broiler chickens were inoculated via eyedrop with live commercial vaccine strains of IBV and NDV alone or in combination to directly evaluate IBV and NDV replication in the trachea at 1, 3, and 5 days after vaccination. Commercial NDV vaccine strains used were B1, VG/GA, and C2. The vaccine strains of IBV tested were Massachusetts (Mass) and Arkansas (Ark). The NDV + Mass vaccines used were commercially manufactured combined products. The NDV + Ark vaccines used were commercial vaccines manufactured as single entity products that were administered by eyedrop to opposite eyes of each chicken.
As measured by qRT-PCR, the replication of NDV strains B1, VG/GA, and C2 did not interfere with the growth of IBV Mass and Ark strain vaccines in the combined vaccine treatment groups. Combination vaccinations using B1 and VG/GA did not interfere with IBV immunity based on challenge or serum antibody production. In the C2 + Mass vaccination trial, IB immunity after challenge was reduced, but it did not seem to be a result of reduced Mass vaccine growth or the ability of the Mass vaccine to induce serum IBV antibody. In contrast, the replication of IBV strains Mass and Ark interfered with the growth of NDV strains B1, VG/GA, and C2 as measured by qRT-PCR. However, interference with NDV replication was not reflected in a reduction in Newcastle disease challenge of immunity findings when combination Mass + NDV products manufactured by vaccine companies were tested. Moreover, NDV immunity was not compromised in two of three trials using single entity vaccines of NDV and Ark
IBV vaccines manufactured separately but administered simultaneously. However, in one trial, NDV immunity was decreased where a NDV single entity product (C2) was given with an IBV single entity Ark vaccine. This finding emphasizes the importance of using manufactured combination vaccines whenever possible to avoid potential interference.
Infectious Bronchitis Virus S1 Gene Sequence Comparison is a Better Predictor of Challenge of Immunity in Chickens than Serotyping by Virus Neutralization
Brian S. Ladman, Alison B. Loupos and Jack Gelb, Jr.
Department of Animal and Food Sciences,
SUMMARY. Five strains of infectious bronchitis virus isolated from commercial chickens from the state of Pennsylvania, USA during the years 1998 and 1999 were studied. The strains were selected for cross-challenge in specific pathogen free chickens and virus neutralization in chick embryos on the basis of partial S1 sequence amino acid identity values. The partial sequences analysed spanned the hypervariable amino terminus region of S1 from amino acid residues 48 to 219, based on the Beaudette strain. Using their S1 identity values, the strains represented a continuum of genetic, and thus antigenic, relationships. When compared with strain PA/5083/99, strain PA/Wolgemuth/98 had high sequence identity (96%) followed by PA/171/99 (85%), PA/5344/98 (70%) and PA/1220/98 (34%). The method of Archetti and Horsfall was used for calculating antigenic relatedness values of virus neutralization tests. The same formula was also applied to the percentage protection values of cross-challenge tests to derive protective relatedness values among the strains. The antigenic relatedness values, protective relatedness values, and the partial S1 sequence identity values were then analysed. The findings indicated partial S1 sequence identity values were more strongly correlated with protective relatedness values and than antigenic relatedness values.
S1 Gene Characteristics and Efficacy of Vaccination against Infectious Bronchitis Virus Field Isolates from the United States and Israel (1996 to 2000)
J. Gelb, Jr.1, Y. Weisman 2, B. S. Ladman1 and R. Meir 2
1Department of Animal and Food Sciences, University of Delaware, Newark, DE 19717, USA, and
SUMMARY. The S1 genes of isolates of avian coronavirus infectious bronchitis virus (IBV) from commercial chickens in the US and Israel (20 isolates from each country) were studied using reverse transcription-polymerase chain reaction restriction fragment length polymorphism and sequencing. Partial sequences spanning the amino terminus region of S1 from amino acid residues 48 to 219, based on the Beaudette strain, were used for analysis. Phylogenetic clustering and high-sequence identity values were used to identify isolates that appeared to be derived from live IBV vaccines used in the two countries. Novel variant strains, unrelated by S1 sequencing and restriction fragment length polymorphism analyses to reference and vaccine strains, were also identified. Based on S1 sequence identity to available vaccines, the potential to use vaccination to control IBV infections was evaluated. Vaccination with commercial live strains Massachusetts (Mass), Arkansas (Ark) or
DE/072/92, generally produced immunity against vaccine-related field isolates displaying high S1 sequence similarities (]/90%) to the respective vaccine strains. Immunization with a bivalent vaccine containing the Mass and Ark strains provided good cross-protection, averaging 81% against challenge with five variant isolates from the US having amino acid identity values ranging from 62 to 69% to Mass and from 68 to 83% to Ark, respectively. In contrast, the H120 vaccine strain induced low levels of protection, ranging from 25 to 58% against variant field isolates from Israel with amino acid identity values from 65 to 67%.