Identification of recent infectious bronchitis virus isolates that are serologically different from current vaccine strains

Three 1986 infectious bronchitis virus (IBV) isolates were serotyped by a hemagglutination-inhibition method and were found to be serologically different from the seven strains currently used in regular and specially licensed vaccines in the United States (Mass 41, H52, H120, Conn 46, Fla 18288, JMK, and Ark 99) and from 13 other reference IBV strains. The recent isolates were from layer flocks that had histories of egg-production declines and shell-quality problems. Two of the isolates, one each from Maine and Ohio, were serologically related to a 1983 IBV isolate from layer replacements in Pennsylvania. The third isolate, from Korea, was serologically different from any of the reference strains and the other new isolates.     

Infectious bronchitis virus serotypes in poultry flocks in Jordan

Infectious bronchitis virus (IBV) causes respiratory disease in chickens all over the world. IBV has many serotypes that do not confer cross protection against each other. Hemagglutination inhibition (HI) test has been used to determine the serotypes of IBV as a substitute to the more laborious virus neutralization test and the more sophisticated restriction endonuclease digestion or sequencing of the S1 gene. In Jordan, no previous studies have been carried out to determine the involvement of IBV in respiratory disease in chickens, or the serotypes of IBV that possibly exist. In this study, serum from different chicken flocks (n = 20) that suffered from respiratory disease were tested for IBV antibodies using commercial IBV antibody ELISA at time of the initial signs of the respiratory disease and repeated on serum samples from the same flocks 10–14 days later. ELISA titer for IBV increased in 14 out of 20 flocks (70%) after 10–14 days of the initial signs of the respiratory disease and this indicates a recent exposure to IBV. The second serum samples from these 14 flocks were further examined against a panel of five IBV antigens (Ark, Conn, DE-072, JMK, and Mass) by HI test to determine the serotype(s) of IBV they have been exposed to. The HI test results indicated that the exposure of some of these flocks were to Ark, DE-072, and Mass like serotypes. However, the HI titers against the antigens used in this study were relatively similar in 10 out of the 14 flocks (71%) and the serotype of IBV that these flocks were exposed to could not be determined and the possible causes of this are discussed.     

Prevalence of Arkansas-type infectious bronchitis virus in Delmarva peninsula chickens

The prevalence of Arkansas (Ark)-type infectious bronchitis virus (IBV) in Delmarva peninsula broiler-type chickens was determined. The immunity of 5-to-11-week-old commercial broilers was evaluated by intraocular inoculation with Ark-type DPI strain (Ark DPI) challenge virus and collection of tracheal swabbings 5 days later. Serum Ark-type antibody titers were obtained using the virus-neutralization test. Eighty-five flocks were tested from January to August 1981. Nearly 60% of the flocks had substantial (greater than or equal to 70%) local immunity of the upper respiratory tract. Twenty-two percent had intermediate (50-69%) and 19% of the flocks had low (less than or equal to 40%) levels of local immunity. Serum antibody titers generally agreed with challenge results. In addition, high Ark-type IBV neutralizing-antibody titers were found in 16 Delmarva broiler breeder flocks. Seven current IBV field isolates were characterized for antigenic similarity to Ark DPI. Four isolates contained Ark antigen(s) based on significant neutralization in virus-neutralization tests and on substantial immunity to challenge afforded by Ark DPI virus immunization. Three isolates did not appear to contain Ark antigen(s). Immunization of chickens with Ark DPI virus afforded substantial protection against Connecticut- and homologous-type virus challenge, partial immunity (63%) against JMK, and no protection against the Massachusetts 41 strain of IBV.     

Molecular and serologic characterization, pathogenicity, and protection studies with infectious bronchitis virus field isolates from California

In this study, we characterized three variant infectious bronchitis virus (IBV) strains isolated in 2003 and 2004 from broiler chickens in California and compared them to previously isolated California variant viruses and to common vaccine serotypes used in the United States. We conducted genetic, serologic, and pathogenicity studies on all three isolates, then tested different vaccines against one of the viruses. Genetically the three variant IBV strains, designated CA557/03, CA706/03, and CA1737/04, were not related to each other. GenBank BLAST database search and phylogenetic analysis of the hypervariable region of the S1 subunit of the spike gene to determine the most closely related viruses to the three variants showed the CA557/03 variant to be 81.8% similar to the CAV/CA56b/91 whereas the CA706/03 and CA1737/04 variant viruses were only distantly related to Dutch/D1466/81 (72.2%), a vaccine strain used in Europe, and Korea/K142/02 (72.7%), a Korean field isolate, respectively. Cross virus-neutralization testing showed that none of the 2003-04 California IBV variant viruses were serologically related to each other or to Ark, Conn, or Mass vaccine strains. In addition the CA1737/04 isolate was also tested against DE072 and found not to be serologically related. All three variant viruses were pathogenic in 1-wk-old broilers and vaccination with Mass/Conn followed by Holland/Conn provided 80% protection against the CA1737/04 virus. The 2003-04 California variant viruses were not compared with variants isolated in California during 1970s and 1980s because, to our knowledge, no genetic information is available and those viruses are no longer obtainable. This study shows that the CA557/03 virus was distantly related to the CAV-type viruses isolated in California in the early 1990s, but that none of the 2003-04 viruses were similar genetically or serologically to the CAL99-type viruses, indicating that new IBV variants continue to emerge and cause disease in commercial chickens in California.     

Detection of avian infectious bronchitis viral infection using in situ hybridization and recombinant DNA

A recombinant DNA probe with specificity for the 3' end of genomic RNA from the Ark 99 strain of infectious bronchitis virus (IBV) was found to hybridize with extracted RNA of three strains with the Ark serotype, as well as the Mass41, Holl52, Gray, JMK, Conn, Fla and SE17 strains of IBV. Viral infection was detected in the cytoplasm of chicken embryo kidney cells inoculated with Mass41, Ark99, SE17 or two recent field isolates of IBV using in situ cytohybridization and a biotinylated probe. In vivo infections were detected in individual cells of tracheas and lungs 2,4, and 6 days after inoculation of chicks with Mass41 and Ark99. In situ hybridization of Ark99 infected tissue sections using 32P-dATP labelled probe indicated that more viral replication was present in the trachea on day 4 than either days 2 or 6; whereas more viral RNA was found in the lungs on day 6 than days 2 or 4 after inoculation.     

Comparison of ciliary activity and virus recovery from tracheas of chickens and humoral immunity after inoculation with serotypes of avian infectious bronchitis virus

Cross-protection tests with homologous and heterologous serotypes of infectious bronchitis virus (IBV) were used to compare ciliary activity and virus recovery from tracheas of chickens. Validation of this technique included correlating the neutralization indices of antiserum obtained from some infected birds. Chickens were inoculated intratracheally with either the JMK or Connecticut (Conn) serotype of IBV. Three weeks later, infected and uninfected groups were challenged by the same route with homologous and heterologous virus. The JMK strain provided immunity against homologous challenge and the Conn strain, as indicated by good ciliary activity and lack of challenge virus recovery. The Conn strain provided only homologous protection, as ciliostasis occurred and virus was recovered after challenge with the JMK strain. In each case, antiserum to immunizing virus neutralized only the homologous virus. Controls were uniformly susceptible and lacked neutralizing antibody. A similar experiment with the Ark 99 serotype and a recent isolate (397) of IBV revealed complete cross-protection of the tracheas. Antiserum to each virus neutralized the homologous and heterologous virus in each case in reciprocal tests. The results indicate that these two viruses are closely related. The complete agreement between ciliary activity and virus isolation indicates that ciliary activity is a reliable, objective criterion upon which tracheal immunity can be judged in cross-protection tests.     

Serological profiles of commercial broiler breeders and their progeny. 1. Infectious bronchitis virus

Serum samples were collected from broiler breeders and their 1-day-old, 2-week-old, and 5-week-old progeny from different regions of the United States. Individual samples were tested by hemagglutination-inhibition (HI) against six infectious bronchitis virus (IBV) strains: Massachusetts 41 (Mass), H52, Connecticut 46 (Conn), Arkansas 99, SE17, and JMK. The use of multiple strains to test broiler flocks resulted in the detection of seroconversions to Conn and JMK vaccination that were not detected with the IBV Mass HI test. Further, HI titers were detected to IBV strains not used for flock vaccination. In some cases, those titers could be due to cross reactions to antigens common to each of the virus strains. In two breeder flocks, the highest HI titers were to heterologous strains.     

鸡传染性支气管炎病毒沈阳分离株生物学特性及其免疫原S1基因的克隆与序列分析

本研究对分离自沈阳地区的一株传染性支气管炎病毒（ＳＹ毒株）进行了生物学特性的研究,同时成功地对其免疫原Ｓ１基因进行了ＲＴ－ＰＣＲ扩增、克隆与序列分析。 通过电镜观察、动物回归试验、血凝特性研究等试验验证分离自沈阳地区的ＳＹ毒株确实为一株传染性支气管炎病毒。气管环组织培养交叉中和试验结果表明,分离株ＳＹ株不同于参考毒株澳大利亚Ｔ、Ｈ５２、Ｍ４１,且不同于国内其它流行株ＨＤ、ＨＢ、ＸＢ、ＤＢ等,是一个新的变异株。 利用ＩＢＶ Ｓ１基因特异性寡聚核苷酸引物,经ＲＴ－ＰＣＲ扩增ＳＹ毒株的Ｓ１基因,得到预期的约１．７Ｋｂ片段;并将扩增所得ｃＤＮＡ插入克隆质粒ｐＵＣ１９的ＥｃｏＲⅠ／ＢａｍＨⅠ位点,在大肠杆菌ＤＨ５ａ中实现目的基因的克隆。经限制性核酸内切酶分析及ＰＣＲ鉴定,证实为阳性重组质粒,利用末端双脱氧链终止法对其测序,得到Ｓ１基因全长１６４０ｂｐ,包括整个开放阅读框。通过序列分析软件ＤＮＡＳＩＳ、ＰＲＯＳＩＳ、ＭＥＧＡ等软件对Ｓ１基因核苷酸序列及推导的氨基酸序列进行分析,结果表明：分离株ＳＹ与７株参考株和国内流行株ＨＤ株相比,无论是核苷酸序列同源百分率还是氨基酸序列同源百分离都较低,均未达到８０％,这就提示我们ＳＹ毒     

Phylogenetic analysis of S1 gene of infectious bronchitis virus isolates from China

Between 2006 and 2009, seven strains of infectious bronchitis (IB) virus (IBV) were isolated from vaccinated chicken flocks on different chicken farms in China. The pathogenic characters of seven IBV strains were assessed. Each of the seven strains was infective to the test chickens and could induce an immune response. The results from chicken embryo cross-neutralization assays showed that these strains were antigenically distinct from classic IBV strains of H120, M41, Conn, and Gray. Compared to H120 vaccine strain, point mutation, short insertion, and deletion occurred at many positions in the S1 protein of the seven strains. Five of the seven strains had the motif (HRRRR), which was identical to that of the epidemic IBV strains in China. Two new motifs (HRLRR and RRIRR) emerged in the isolated strains. The homology of the nucleotide and amino acid sequences of the S1 gene among the seven isolates was 81.7%-99.7% and 79.0%-99.4%, respectively. These seven strains were also genetically different from the vaccine strains and non-China IBV strains but closely related to large numbers of Chinese strains. The seven isolates and 36 reference IBV strains were clustered into six distinct groups (I-VI). The seven strains were categorized into groups I, II, and III, forming a big phylogenetic branch, which is closely related to Chinese IBVs, whereas the vaccine strains belonging to group VI are genetically distant from groups I, II, and III. The results from this study indicate that different IBV strains cocirculate in the chicken population in China.     

Evaluation of the hemagglutination-inhibition test for measuring the response of chickens to avian infectious bronchitis virus vaccination

An infectious bronchitis virus (IBV) hemagglutination-inhibition (HI) test was used to assay serum-antibody titers after IBV vaccination of IBV-susceptible specific-pathogen-free broilers and commercial layers. Three-week-old broilers were vaccinated via eye-drop with IBV strains that represent the antigenic spectrum of commercial vaccines--Holland, Massachusetts 41 (41 Ms), Connecticut 46, Florida 18288, or JMK strain--and revaccinated 3 weeks later with either the same or a heterologous strain. Weekly serum samples were tested by IBV HI with homologous and heterologous antigens. Vaccinates, except for those vaccinated with the Holland strain, were HI-positive with homologous but not heterologous antigens by 1 to 2 weeks postvaccination. Sixteen-week-old IBV-vaccinated commercial layers were revaccinated with IBV Holland 52 (H 52) strain and subsequently infected with Arkansas 99 (Ark 99) and SE 17 strains. In contrast to the limited HI cross-reactivity of serum from IBV-vaccinated broilers, there were extensive cross-reactions in HI tests with 41 Ms, H 52, Ark 99, and SE 17 antigens of revaccinated layers. These results demonstrate that the IBV HI test is more strain-specific than previous reports indicate, especially when the test samples are from early postvaccination.     

Protection afforded infectious bronchitis virus-vaccinated sentinel chickens raised in a commercial environment

The protective efficacy of three infectious bronchitis virus (IBV) vaccines for sentinel chickens raised with commercial Delmarva broiler chickens was evaluated during winter 1987. Specific-pathogen-free leghorn sentinel chickens were vaccinated with Massachusetts (Mass) alone, Mass and JMK, or Mass and Arkansas (Ark) combination live vaccines, or they remained unvaccinated. Four weeks post-vaccination, sentinels were placed on broiler farms at weekly intervals for 3 weeks corresponding to weeks 4, 5, and 6 of the broiler growing cycle. Vaccine efficacy was evaluated based on IBV reisolation attempts from tracheal swabbings following a 1-week field exposure period. Sentinel chickens vaccinated with Mass and Ark combination vaccine were best protected against IBV field challenge. Only four IBV isolations were made out of a 3-week total of 36 attempts, for an 11% isolation rate. IBV vaccines containing either Mass alone or Mass and JMK offered much lower levels of protection.     

鸡传染性支气管炎病毒我国地方分离株S1基因的分子特征

应用反转录 -聚合酶链式反应 ,以 IBVS1基因的特异性引物从新疆 IBV流行株基因组中扩增出预期的 1 .7kb左右的片段 ,将此PCR产物修饰后插入到克隆质粒 p UC1 9的Sma I位点 ,在大肠杆菌中实现了目的基因的分子克隆。经测序及序列比较 ,与国外参考毒株相比表明新疆 IBV分离株已有分子水平的变异 ,与国内分离株 QX同源性高达 96% ,证明这两个分离株有很高的同源性     

2株鸡传染性支气管炎病毒的分离鉴定及S1基因的序列分析

采用RT-PCR方法对2株传染性支气管炎病毒分离株的S1基因进行序列扩增、测定及分析,应用DNAStar软件将这些分离毒株与中国常用疫苗毒株、其他血清型的代表参考毒株分别进行核苷酸和氨基酸系统发生进化关系分析。核苷酸与氨基酸序列分析结果均表明：2株分离毒株与J2、Q1、T3毒株的亲缘关系较近,与疫苗株H120和4/91的亲缘关系较远。     

鸡肾型传染性支气管炎病毒分离株S1基因的序列分析

采取RT-PCR方法对5株传染性支气管炎病毒分离株的S1基因进行序列扩增、测定及分析,应用DNAStar软件将这些分离毒株与中国常用疫苗毒株、国际上其它血清型的代表参考毒株分别进行核苷酸和氨基酸系统发生进化关系分析。核苷酸与氨基酸序列分析结果表明,这5株分离毒株与A2毒株的亲缘关系较近,与疫苗株H120和Ma5的亲缘关系较远。