Emergence of Subtype Strains of the Arkansas Serotype of Infectious Bronchitis Virus in Delmarva Broiler Chickens

Abstract

Infectious bronchitis virus (IBV) field isolates of the Arkansas (Ark) serotype were identified by reverse transcription-polymerase chain reaction (RT-PCR) as the most common serotype isolated from 1993 to 1997. These isolates were recovered from broiler flocks with respiratory disease raised on the Delmarva peninsula in spite of Ark vaccination in the region. For the purposes of investigating this apparently paradoxical finding, five RT-PCR Ark-positive field isolates recovered in 1995 and 1996 were selected for further characterization. The isolates were compared with Ark reference strains by reciprocal virus neutralization (VN) in embryonated eggs, S-1 gene sequence analysis, and challenge of immunity studies in specific-pathogen-free (SPF) chickens. Antigenic (VN) comparisons and S-1 gene analysis confirmed that the five RT-PCR Ark-positive field isolates were of the Ark serotype but also revealed that the viruses could be readily distinguished from Ark reference strains. Four of the isolates (Ark/213/96, Ark/15C/96, Ark/1529/95, Ark/1534/95) were found to have higher antigenic relatedness percentages to each other (95%-100%) than to Ark reference strains DPI (52%-72%) and Georgia variant (Georgia var) (53%-68%) by VN. Another isolate, Ark/1535/95, was found to differ antigenically from the other four RT-PCR Ark-positive field isolates (34%-61%), Ark DPI (44%), and Georgia var (43%) strains. The trends in the S-1 gene sequencing results were similar to those observed for the VN findings. Isolates Ark/213/96, Ark/15C/96, Ark/1529/95, and Ark/1534/95 demonstrated a higher degree of predicted S-1 amino acid similarity to each other (96.5%-98.7%) than to Ark DPI (92.4%-93.7%), Ark 99 (93.2%-94.7%), and Georgia var (89.3%-90.8%). Ark/1535/95 S-1 amino acid similarity values were lower compared with those of the other four RT-PCR Ark-positive field isolates (93.4%-94.8%), Ark DPI (91.9%), Ark 99 (93.0%), and Georgia var (88.7%). Furthermore, the isolates could be distinguished from the Ark reference strains by a characteristic sequence polymorphism, a six-nucleotide deletion encoding amino acids 57 (Asp) and 58 (Asp) in hypervariable region 1 of S-1. On the basis of the VN and sequencing findings, isolates Ark/213/96, Ark/15C/96, Ark/1529/95, and Ark/1534/95 were considered to be a single subtype of the Ark serotype. The fifth isolate, Ark/1535/95, may constitute another subtype of the Ark serotype. Vaccination of SPF chickens with a high-titering commercially available live vaccine containing the Ark DPI strain provided solid protection (>90%) against challenge with the RT-PCR Ark-positive field isolates. Immunization of SPF chickens with Ark/213/96 produced 100% protection against challenge with the homologous strain, as well as isolates Ark/1535/95 and Ark 99 but lower levels of protection against Ark DPI (58%) and Georgia var (55%). Primers for RT-PCR were designed to distinguish between the Ark subtypes and the Ark reference strains on the basis of the characteristic six-nucleotide deletion identified in the S...

Rapid selection in chickens of subpopulations within ArkDPI-derived infectious bronchitis virus vaccines

“…In contrast, Nix et al (2000) reported an increasing frequency of Arkansas (Ark)-type IBV isolations in the Delmarva Peninsula (northeastern United States) between 1992 and 1997 in spite of use of ArkDPI-derived vaccines. In the southeastern United States, over one-half of IBV isolates obtained between 1997 and 2002 were ArkDPI-like (Toro et al, 2006).…”

“…At least four different commercial Ark-serotype vaccines derived from this attenuated stock are used in the United States. Two different commercial Ark vaccines, as well as the attenuated ArkDPI seed stock distributed to vaccine companies, and the original unattenuated ArkDPI isolate, have been shown to contain a minor subpopulation similar to an Ark subtype isolated from broilers in the Delmarva Peninsula (Nix et al, 2000). This minor subpopulation was identified by restriction fragment length polymorphism (RFLP) analysis of the S1 gene, real-time polymerase chain reaction (RT-PCR) using primers specific for the subpopulation, and sequencing of the RT-PCR product, but the sequence of the entire S1 gene of the putative subpopulation was not determined.…”

“…The authors of the study suggested that the attenuated Ark DPI vaccines were a ''likely source'' of the pathogenic Ark subtype. They pointed out that neither the original virulent ArkDPI isolate nor its attenuated derivative had been cloned by limiting dilution (Nix et al, 2000). They consider heterogeneity of vaccines desirable, because the viral subpopulations would make the vaccines more antigenically diverse and thus more broadly protective.…”

Combined infectious bronchitis virus Arkansas and Massachusetts serotype vaccination suppresses replication of Arkansas vaccine virus

“…Effective control of infectious bronchitis virus (IBV) has been distinctively difficult because of genetic diversity and continuing evolution of the causal coronavirus (Toro et al, 2012b). In the United States, Arkansas (Ark) type strains have been the most frequently identified type of IBV ranging from 23% to 65% of total isolations per year (Nix et al, 2000;Jackwood et al, 2005;Toro et al, 2006) in spite of Ark serotype vaccination. Other IBV serotypes determined to be widespread in the United States include the Connecticut (Conn), DE072, Massachusetts (Mass) and Georgia serotypes.…”

Genetic Analysis of the S1 Gene of 4/91 Type Infectious Bronchitis Virus Isolated in Japan

“…Various serological types of live or inactivated vaccines have been available for control of IBV infections in Japan. Vaccines have generally been effective in controlling clinical disease associated with IBV infections, but new variants may continue to emerge and cause clinical disease and production problems in vaccinated flocks [8,18]. Some studies [7,10] have reported frequent isolation of isolates related to vaccine strains from diseased flocks in the field, and reversion of the virulence of vaccine strains and recombination has been suspected.…”