Isolation and identification of four infectious bronchitis virus strains in China and analyses of their S1 glycoprotein gene

Between 2003 and 2005, four strains of infectious bronchitis virus (IBV) were isolated from the vaccinated chicken flocks in China. The results from chicken embryo cross-neutralization assays showed that all the four isolates were relative to strain A2 of IBV, which was isolated in 1996 in Beijing and related to strain 4/91. The S1 gene of the spike protein was amplified and sequenced. The nucleotide and amino acid sequence of the S1 gene had a similar degree of identity (88.98-99.28%) among the four Chinese IBV isolates. The identity of the S1 protein gene between the four Chinese IBV isolates and 14 strains of other IBVs varied from 70.06 to 81.59%. Phylogenetic analysis suggested that there are at least four groups of IBVs circulating in China and the disease outbreaks might have been caused by infection of multiple strains of IBV.     

Occurrence and significance of infectious bronchitis virus variant strains in egg and broiler production in the Netherlands

Despite vaccination against Infectious bronchitis virus (IBV) with the Massachusetts type vaccine viruses H120 and H52 in the Netherlands, an increasing number of properly vaccinated flocks have suffered from the disease since 1978. In the years 1978-1982, the virus was isolated from 162 IBV suspected flocks. Cross-virus-neutralization tests showed that the majority (67 per cent) of these isolates belonged to serotypes other than the Massachusetts type, the Connecticut-, Florida-, Iowa 97-, Iowa 609- and JMK serotype. The majority of these Dutch isolates could be divided into 4 serogroups, called D207, D212, D3128 and D3896. Only a few isolates were not related to these serotypes. A survey of 328 flocks for antibody against these serotypes demonstrated that antibody against one or more of these novel serotypes were present in most of the flocks. Experiments demonstrated that vaccination with the H120 vaccine virus was not able to protect chickens against the adverse effects of a challenge with the novel serotypes.     

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.     

First evidence of the emergence of novel putative infectious bronchitis virus genotypes in Cuba

The emergence of new infectious bronchitis virus (IBV) genotypes or serotypes along with the poor cross-protection observed among IBV serotypes have complicated the avian infectious bronchitis (IB) control programs in different geographic regions. In Cuba, the lack of genetic information regarding IBV and the increasing epidemiological importance of this virus in Cuban chicken flocks demand further characterization of IBV isolates. In the present work, studies of genetic diversity and phylogenetic relationships among recent IBV isolates from Cuban chicken flocks showing respiratory disorders were performed. Two putative genotypes genetically different to the Massachusetts genotype H120 strain used in the Cuban vaccination program were found in the flocks assessed. In addition, a potential nephropathogenic IBV isolate was found by first time in Cuba.     

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.     

鸡传染性支气管炎病毒地方流行株的分离与鉴定

从山西各地区疑似鸡传染性支气管炎(IB)的病料中,分离到5株鸡传染性支气管炎病毒(IBV)分离株,并对分离病毒进行了病毒形态观察、对鸡新城疫病毒(NDV)的干扰、鸡胚致病性试验、动物回归试验、血凝特性试验、病毒理化特性测定等生物特性鉴定及IBV N基因特异性片段的检测.电镜观察,可见直径为60～120 am,有囊膜及纤突呈冠状排列的病毒粒子;对NDV有明显的干扰作用;分离株的传代物均有明显的致鸡胚矮小化作用;动物回归感染死亡鸡肾脏病变明显,表现肾脏肿大、花斑肾现象,输尿管内充塞大量尿酸盐;无直接血凝性,经1%胰酶处理后可凝集鸡红细胞;分离株对乙醚和氯仿敏感;采用反转录-聚合酶链式反应(RT-PCR)对分离毒株进行扩增,结果均扩增出特异N基因核酸片段.     

Antigenicity of two turkey astrovirus isolates

Astroviruses are positive-sense single-stranded RNA viruses. These viruses cause gastroenteritis in humans and in a variety of animal species, including turkey poults. Only human astroviruses are well characterized antigenically. In the current study, two turkey astrovirus isolates, TAstV1987 and TAstV2001, were antigenically compared using cross-neutralization tests in turkey embryos, as well as cross-reactivity of the two isolates by an enzyme-linked immunosorbent assay (ELISA). The antigenic relatedness values (R) were calculated using the Archetti and Horsfall formula. The R value based on the cross-neutralization tests was 0.56%, which indicates that TAstV1987 and TAstV2001 belong to different serotypes; the R value of the two viruses based on ELISA was 70.7%, which suggests these two viruses share common antigen(s).     

Genetic diversity of avian infectious bronchitis coronavirus in recent years in China

Fifty-six isolates of avian infectious bronchitis virus (IBV) were obtained from different field outbreaks in China in 2010, and they were genotyped by comparison with 19 reference strains in the present study. The results showed that LX4-type isolates are still the predominant IBVs circulating in chicken flocks in China, and these isolates could be grouped further into two clusters. Viruses in each cluster had favored amino acid residues at different positions in the S1 subunit of the spike protein. In addition, a recombination event was observed to have occurred between LX4- and tl/CH/LDT3/03I-type strains, which contributed to the emergence of a new strain. The most important finding of the study is the isolation and identification of Taiwan II-type (TW II-type) strains of IBV in mainland China in recent years. The genome of TW II-type IBV strains isolated in mainland China has experienced mutations and deletions, as demonstrated by comparison of the entire genome sequence with those of IBV strains isolated in Taiwan. Pathogenicity testing and sequence analysis of the 3' terminal untranslated region revealed that TW II-type IBV strains isolated in mainland China have a close relationship with the embryo-passaged, attenuated TW2296/95.     

Vaccine efficacy against Ontario isolates of infectious bronchitis virus

Infectious bronchitis (IB) is an economically important viral disease with worldwide distribution. Every country with an intensive poultry industry has infectious bronchitis virus (IBV). The virus rapidly spreads from bird to bird through horizontal transmission by aerosol or ingestion. Sentinel bird studies were carried out in southern Ontario and IBV has been isolated from layer flocks. Genetic analysis of the S1 region of the strains showed that they were not vaccine related. The pathogenicity of selected Ontario variants of IBV isolates was studied and the subsequent work was to determine the degree of protection against field isolates provided by a commonly used vaccine MILDVAC-Ma5 in Ontario. The protection was evaluated by challenging immunized chickens with the respiratory (IBV-ON1) and nephropathogenic (IBV-ON4) viruses. The mean vaccine efficacy for IBV-ON1 was 66.7% indicating that a Massachusetts serotype vaccine would provide some protection against IBV field isolates.     

Characterization of infectious bronchitis viruses isolated from outbreaks of disease in commercial flocks in Brazil

Fifteen isolations of infectious bronchitis (IB) virus were made from a total of 126 Brazilian poultry flocks of all ages that were examined. These flocks (14 chicken and 1 quail) were experiencing a variety of IB-like conditions including respiratory disease, digestive and kidney problems, and drops in egg production. One of the isolates was of the Massachusetts serotype. The remainder were examined by means of cross-neutralization tests in tracheal organ cultures and were shown to belong to at least four antigenic groups, all different from ones described previously in other countries. Some, but not all, of the flocks from which they were isolated had been vaccinated against IB with vaccines of the Massachusetts serotype. In vivo protection studies showed that the MA5 vaccine (of the Massachusetts serotype) protected well against challenge with four of these isolates, representing the different serotypes reported in this study.     

Characterisation of avian adenoviruses associated with inclusion body hepatitis

Eleven avian adenoviruses were isolated in monolayer cultures of specific pathogen free chicken kidney cells which were inoculated with suspensions of liver, intestine or bursa obtained from 15 broiler flocks experiencing outbreaks of inclusion body hepatitis (10 isolates) and from five unaffected flocks (one isolate). Of the 11 isolates obtained, nine were identified by virus neutralisation tests as serotype 8, one as serotype 1 and one as serotype 12. Adeno-associated viruses were only observed in combination with adenoviral particles of the serotype 12 isolate which was derived from a relatively mild outbreak of inclusion body hepatitis. Only the serotype 1 isolate, obtained from the unaffected broiler flock, consistently caused the death of embryos with marked pathological changes. All of the isolates produced basophilic intranuclear inclusion bodies surrounded by clear halos in chicken kidney cell cultures. DNA preparations, obtained from six strains of serotype 8 avian adenovirus (two New Zealand isolates, three Australian isolates and the reference strain HVI) after digestion with the restriction enzymes EcoRI and BamHI, gave electrophoretic patterns showing the New Zealand isolates to be similar to one another and to strain HVI, but quite distinct from the Australian isolates.     

Rapid serological profiling by enzyme-linked immunosorbent assay. IV. Association of infectious bursal disease serology with broiler flock performance

Breeder and broiler flocks were serologically evaluated using a multiple enzyme-linked immunosorbent assay (M-ELISA). The serologic status of two commercial broiler-breeder flocks and their progeny was monitored, and 840 sera were promptly assessed for antibodies against six infectious agents using the M-ELISA. Breeder flocks were sampled at lay, and broiler chicks were hatched from fertile eggs collected on the scheduled lay date of the breeders. The broiler chicks were placed for growout as eight separate flocks (four from each breeder), and the serologic survey of broilers included sequentially sampling each flock five times between 1 day of age and market. Association of broiler vaccination schedules, mortality, and condemnation data with the temporal serologic data obtained indicated that the earlier appearance of active antibody against infectious bursal disease (IBD) in some unvaccinated flocks was associated with subsequent higher growout mortality and with the poorer overall performance that these flocks experienced. The results of this serologic survey also demonstrated that if a constant, well-timed monitoring program had not been used, major serologic differences between flocks would not have been detected. Serologic profiles of selected broiler flocks by virus-neutralization (VN) tests for infectious bronchitis virus (IBV) and reovirus or by hemagglutination-inhibition (HI) tests for Newcastle disease virus (NDV) compared favorably with the serologic profiles obtained by M-ELISA. Comparison of vaccination histories with serologic results derived from M-ELISA, VN or HI tests indicated that response to vaccination for IBV and NDV at 1 day was either blocked or significantly delayed by moderate levels of maternal antibody and/or were suppressed by an apparent field outbreak of IBD that occurred in all eight broiler flocks.     

Detection and molecular characterization of infectious bronchitis virus isolated from recent outbreaks in broiler flocks in Thailand

Thirteen field isolates of infectious bronchitis virus (IBV) were isolated from broiler flocks in Thailand between January and June 2008. The 878-bp of the S1 gene covering a hypervariable region was amplified and sequenced. Phylogenetic analysis based on that region revealed that these viruses were separated into two groups (I and II). IBV isolates in group I were not related to other IBV strains published in the GenBank database. Group 1 nucleotide sequence identities were less than 85% and amino acid sequence identities less than 84% in common with IBVs published in the GenBank database. This group likely represents the strains indigenous to Thailand. The isolates in group II showed a close relationship with Chinese IBVs. They had nucleotide sequence identities of 97-98% and amino acid sequence identities 96-98% in common with Chinese IBVs (strain A2, SH and QXIBV). This finding indicated that the recent Thai IBVs evolved separately and at least two groups of viruses are circulating in Thailand.     

Genetic diversity of avian infectious bronchitis virus isolates in Korea between 2003 and 2006

Thirty-three field isolates of avian infectious bronchitis virus (IBV) were recovered from commercial chicken flocks in Korea between 2003 and 2006 and were characterized phylogenetically by nucleotide sequence analysis of the IBV S1 gene hyper-variable region. Our phylogenetic analysis revealed that recent field isolates of IBV formed at least three distinct phylogenetic types, including K-I, K-II, and K-III. K-I type IBV consisted of indigenous, 13 IBV isolates which evolved from the Kr-EJ/95 strain and then separated into the lineages of type K-Ia and type K-Ib. K-II type IBV isolates (n = 19) were closely related to nephropathogenic IBV variants from China and Japan. The K-III type isolate (Kr/D064/05), first identified by this study, was closely related to enteric IBV variants from the Chinese strains that cause proventriculitis. Sequence comparisons showed amino acid differences of >27.5% between IBV types. The molecular epidemiologic characteristics of IBV field isolates are briefly discussed.     

Viral pathogenesis of a nephrotropic infectious bronchitis virus isolated from commercial pullets.

Infectious bronchitis was diagnosed in 3-to-4-week-old pullets from an outbreak in a commercial flock in California. The disease was characterized by head swelling, watery discharge from the eyes and nostrils, and urates in kidneys. Mortality ranged from 1.8% to 12.5% per week. The isolation of a coronavirus from a suspension of pooled kidneys from clinically ill chickens at the fifth passage in 10-day-old chicken embryos, gross and histologic renal lesions, and seroconversion by enzyme-linked immunosorbent assay in inoculated birds suggested that the virus isolated was a nephrotrophic strain of infectious bronchitis virus (IBV). The virus isolate was found to be a previously unrecognized serotype, based on virus neutralization tests performed in embryonated chicken eggs. Nephropathogenicity of the IBV isolate was confirmed by inoculation of the viral isolate into specific-pathogen-free chicks and demonstration of renal lesions. The isolation of nephrotropic strains of IBV has not been reported previously from poultry in California.     

Egg production in relation to the results of a long term serological survey of 73 flocks of fowl

Summary Seventy-three flocks of fowl were tested at regular intervals for the presence of precipitins to fowl adenovirus (AV) and infectious bronchitis virus (IBV), haem-agglutinating inhibiting antibodies to BC14 virus, and of agglutinins to Mycoplasma gallisepticum (M.g.) and Mycoplasma synoviae (M.s.). In all the eight flocks affected with Egg Drop Syndrome (EDS '76), egg production problems were associated with increasing numbers of BCI4 virus reactors and AV reactors. In flocks showing production problems other than EDS '76 without any apparent cause, the average percentage of AV reactors increased significantly after the rearing period; this was not true of IBV reactors. BC14 reactors were either absent or present only once, in small numbers and with low titres, during the test period. The average percentage of AV reactors did not increase after the rearing period either in normally producing flocks or in flocks with production problems for which other diseases or dietary errors plausibly accounted for these problems. All this suggests a pathogenic role of AV in production problems. One can conclude from the high percentage of reactors in all groups of flocks that sub-clinical IBV infections are common. The percentage of IBV reactors during the laying period of flocks with EDS '76 was significantly higher than that of normally producing flocks. It is therefore suggested that subclinical IBV infection could be among the factors causing stress, acting as a trigger for EDS '76. All M.g.-infected flocks showed production problems; M.s. infections could not be related to egg production disturbances.     

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.