1
Variation in the spike protein of
the 793/B type of infectious
bronchitis virus, in the field and
during alternate passage in
chickens and embryonated eggs
Authors: David Cavanagh a; Jean-Paul Picault b; Richard E. Gough c; Michael Hess d; Karen
Mawditt a; Paul Britton a
Affiliations:
a
Compton Laboratory, Institute for Animal Health. Compton, Newbury. UK
Ploufragan Laboratory, French Food Safety Agency (AFSSA). Ploufragan. France
c
Avian Virology, Veterinary Laboratories Agency Weybridge. Addlestone, Surrey.
UK
d
Intervet UK Ltd. Houghton, Huntingdon. UK
b
DOI: 10.1080/03079450400025414
Publication Frequency: 6 issues per year
Published in:
Avian Pathology, Volume 34, Issue 1 February 2005 , pages 20 - 25
Abstract
The degree of variation exhibited within the 793/B serotype (also known as 4/91 and CR88 serotypes)
was investigated with nine French and 10 British isolates, collected between 1985 and 1994. The S1
part (1644 nucleotides) of the spike protein gene of the first known isolate of this serotype,
FR/CR85131/85, had 95.9% to 97% nucleotide identity with the other isolates. Partial sequencing of
isolates from Iran and Saudi Arabia, isolated in 2000, revealed approximately 95% nucleotide identity
with European isolates, including the two live 793/B vaccinal strains, showing that they were not reisolations of vaccinal virus. The data indicates that strains within the 793/B serotype have 96%
nucleotide identity within the whole S1 gene and 93% nucleotide identity within the first 560
nucleotides, and 92% and 86% amino acid identities in the corresponding protein regions. This is
similar to the identities exhibited within the Massachusetts serotype.
Sequence analysis of a 793/B field isolate after passage in embryonated eggs, then in chickens and
then again in eggs revealed selection for a serine and alanine at S1 amino acid position 95 in chickenpassaged and egg-passaged virus, respectively. There was no change in pathogenicity. This is the first
demonstration at gene sequence level of host-driven selection for infectious bronchitis virus.
2
Introduction
Infectious bronchitis virus (IBV), together with Turkey coronavirus (Guy, 2000; Cavanagh, 2001a,b;
Cavanagh et al., 2001) and Pheasant coronavirus (Cavanagh et al., 2002; Welchman et al., 2002),
form Group 3 within the genus Coronavirus, family Coronaviridae (Cavanagh, 1997; Cavanagh,
2001a; Enjuanes et al., 2000). Although usually associated with respiratory disease (Matthijs et al.,
2003) and drops in egg production, some strains also cause nephritis (Cook et al., 2001b; Li & Yang,
2001).
It has been known for many years that IBV exists as dozens of serotypes (Cavanagh & Naqi, 2003).
The serotype is defined by the virus neutralization test, the defining antibodies being against the
amino-terminal half, S1, of the large spike glycoprotein (S) (Koch et al., 1990; Kant et al., 1992). In
keeping with the large number of IBV serotypes, the S1 protein is very variable; serotypes commonly
differ by 20% to 25%, and up to 50%, of amino acids. There have been few in-depth analyses of the
degree of variation within IBV serotypes (Cavanagh et al., 1988, 1992; Lee & Jackwood, 2001). In
this paper we report the intra-serotype variation exhibited by the 793/B serotype (Gough et al., 1992),
also known as 4/91 (Parsons et al., 1992) and CR88 (Picault et al., 1995). Virus of this serotype has
been detected in several European countries, although at different frequencies (Cook et al., 1996;
Capua et al., 1999; Cavanagh et al., 1999; Picault et al., 1995; Meulemans et al., 2001), although not
in some others (Farsang et al., 2002). This serotype may have entered the UK in the winter of 1990/91,
when it was sometimes associated with deep pectoral muscle myopathy in layers, in addition to the
more usual manifestations of infectious bronchitis (Gough et al., 1992; Parsons et al., 1992).
Subsequently it was discovered that this serotype had been present in France for several years. Indeed,
the first known strain of this serotype was isolated there in 1985 (Picault et al., 1995).
Jackwood and colleagues have compared the S1 gene sequence of a 793/B-type field virus and its eggpassaged derivative and reported a difference of amino acid at residue 95 (Callison et al., 2001). We
have investigated this further to ascertain whether this difference is host-associated.
Materials and methods
Isolates of the 793/B serotype
The nomenclature of the isolates used here is that proposed by Cavanagh (2001a): country/isolate
number/year of isolation. The names of the countries of origin in this report have been abbreviated
thus: France (FR), Iran (IR), Saudi Arabia (SA) and United Kingdom (UK). The UK isolates isolated
in 1991 and 1993 have been described by Adzhar et al. (1997). These and the names of the other
isolates investigated are shown later ('Databank gene sequence accession numbers').
Extraction of RNA
RNA was extracted as described previously, using guanidinium isothiocyanate, and dissolved in 20 l
water (Cavanagh et al., 2002).
3
Reverse transcription-polymerase chain reaction
The reverse transcription-polymerase chain reactions (RT-PCRs) were performed as previously
described (Cook et al., 2001a; Cavanagh et al., 2002). The whole of the S1 part of the S gene was
amplified using oligonucleotides IBP1- and S1Uni2+ (Adzhar et al., 1996, 1997). Nucleotides 92 to
570 of S1 were amplified using oligonucleotides B1+ (92 5'-AAAGTGCCTTTAGGCCTGG-3' 110)
and B88- (557 5'-GCCCACGTCCGCAAA-3' 570), where the numbers refer to the positions of the
first and last nucleotides in relation to the S1 gene sequence of 793/B-type isolates (Adzhar et al.,
1997). Oligonucleotides B1 and B88 were designed to be specific to the 793/B serotype. The more
general oligonucleotides XCE1- and XCE2+ (Capua et al., 1999; Cavanagh et al., 1999),
corresponding to nucleotides 709 to 1166 in the S1 gene of the 793/B serotype, were used to amplify
part of S1 of the Iranian and Saudi Arabian isolates.
Nucleotide sequencing
Sequencing of PCR products was performed using either the Sequenase PCR product sequencing kit
(USB; Amersham, Little Chalfont, Bucks., UK) or the Thermosequenase dye terminator cycle
sequencing kit, version 2.0 (Amersham). The PCR products complementary to the whole of S1 were
sequenced using a number of oligonucleotides as sequencing primers (Adzhar et al., 1997) while the
smaller PCR products were sequenced using the same oligonucleotides as had been used to amplify
them.
Sequence analysis
Sequences were aligned using ClustalX version 1.64b (Thompson et al., 1994) and compared using
GeneDoc Multiple Sequence Alignment Editor and Shading Utility version 2.5.000
(http://www.psc.edu/biomed/genedoc). Aligned sequences were analysed using PHYLIP version
3.57C (http://evolution.genetics.washington.edu/phylip.html; Felsentein, 1993). The trees were
constructed using the neighbour-joining programme and viewed using Treeview
(http://taxonomy.zoology.gla.ac.uk/fod/treeview.html).
Databank gene sequence accession numbers
First region in S1 (up to nucleotide 561): FR/CR85131/85 (AJ619606); FR/CR88061/88
(AJ619607); FR/CR88099/88 (AJ619608); FR/CR88121/88 (AJ619609); FR/CR94007/94
(AJ619610); FR/CR94047/94 (AJ619611); FR/CR94048/94 (AJ619612); FR/CR94247/94
(AJ619613); FR/CR94279/94 (AJ619614); FR/CR94364/94 (AJ619615); UK/1/91 (AJ619616);
UK/2/91 (AJ619617); UK/3/91 (AJ619618); UK/5/91 (AJ619619); UK/6/91 (AJ619620); UK/7/93
(AJ619621); UK/8/93 (AJ619622); UK/9/93 (AJ619623); UK/10/93 (AJ619624); UK/11/93
(AJ619625); UK/12/93 (AJ619626); UK/42/96 (AJ619627); UK/159/93 (AJ619628); UK/186/96
(AJ619629); UK/236/95 (AJ619630); UK/574/96 (AJ619631); UK/1233/95 (AJ619632).
Complete S1 sequences: UK/1233/95 (AJ618984); FR/CR85131/85 (AJ618985); FR/CR88061/88
(AJ618986); FR/CR94047/94 (AJ618987). Previously published complete S1 sequences, included in
the analyses: UK/7/91 (Z83975); UK/2/91 (Z83976); UK/3/91 (Z83977); UK/5/91 (Z83978); UK/7/93
(Z83979).
4
Results
Mid-1990s UK isolates were very similar to earlier 793/B-type strains
Each isolate that we analysed was probably a quasispecies (i.e. there would have been heterogeneity
at some nucleotide positions). Our sequencing approach was to sequence PCR products. The sequence
obtained reflects the nucleotides that were possessed by the majority of the viral RNA molecules at
each nucleotide position. All the British and French strains in this study were isolated before vaccines
of the 793/B serotype were used in the field.
Following sequencing of nucleotides 92-557 of the S1 gene, the UK isolates of the 793/B serotype
isolated between 1991 and 1996 had nucleotide identity of 94.1%, compared with approximately 70%
identity with many other serotypes.
Sequence analysis of French isolates of the 793/B serotype
Virus neutralization tests in the mid-1990s had shown that many IBVs isolated in France since 1988
were serologically related to the 793/B-type isolates of the UK (Picault et al., 1995). Our retrospective
analysis has further revealed that another 793/B-serotype French isolate, FR/CR85131/85, isolated in
1985, is the oldest known isolate of this serotype. There is no evidence to suggest that this isolate was
the progenitor of the others. Notwithstanding, FR/CR85131/85 had 95.3% to 97.4% nucleotide identity
with the other French isolates.
Comparison of the French and British isolates
Clustal analysis of the first 560 or so S1 nucleotides of the French and British isolates showed that
most, but not all, of them tended to segregate along geographical lines (Figure 1a). A similar, although
not identical, distribution was evident when the amino acid sequences were compared (Figure 1b). The
degree of identity of FR/CR85131/85 with the British isolates ( 94.2%) was essentially the same as
that among the French isolates. This is reflected not only in the Clustal diagram based on the first 560
nucleotides (Figure 1), but also that based on complete S1 sequences (data not shown).
5
Figure 1. Relationships between the UK and French isolates of the 793/B serotype illustrated by a
maximum likelihood phylogeny unrooted tree, based on nucleotides 1 to 561. (a) Nucleotide
sequences. (b) Amino acid sequences. The French isolates are shown in bold. The oldest isolate,
FR/CR85131/85, has been boxed.
6
The results show that within the 793/B serotype, nucleotide identity in the first 560 nucleotides was
93%, compared with about 70% when one compares the nucleotides of most serotypes with other
serotypes. At the deduced amino acid level, FR/CR85131/85 had 86.5% identity with the French and
British isolates in the first 560 nucleotides, compared with approximately 65% when comparing
different serotypes.
The S1 genes of three of the French isolates, FR/CR85131/85, FR/CR88061/88 and FR/CR94047/94,
were sequenced in their entirety. The S1 gene of the 1985 French isolate had 95.9% and 97.0%
nucleotide identity with the other two French isolates, respectively, and 96.1% to 96.9% identity with
the five UK isolates for which the complete S1 gene sequence had been established. Thus, nucleotide
identity within the whole of the S1 gene for 793/B-serotype isolates was 96%, compared with
generally 80% between many serotypes. The deduced amino acid sequences of the complete S1
proteins had 92.4% identity with the oldest known 793/B isolate, FR/CR85131/85, compared with
75% to 80% between many serotypes.
Synonymous (s) and non-synonymous (ns) genetic changes are those that do not and do, respectively,
cause amino acid differences. The average ratio ds/dns for the whole S1 sequences of the 793/B
serotype was 1.7. The ds/dns ratios following comparisons of whole S1 sequences of representative
strains of different serotypes (Massachusetts M41 strain, D274 strain D207 and 793/B strain
CR85131/85) were in the range 3.4 to 5.5 (mean, 4.4).
Sequence analysis of isolates from Iran and Saudi Arabia
Two isolates from Iran and one from Saudi Arabia, isolated in 2000, had been characterized as being
of the 793/B serotype by the virus neutralization test. Corroborative evidence was sought by
sequencing S1 nucleotides 802 to 1081 after RT-PCR using general IBV oligonucleotide primers
XCE1- and XCE2+ (Capua et al., 1999; Cavanagh et al., 1999). The two Iranian isolates had 97.5%
identity with each other, and had 94.7% and 96.4% nucleotide identity, respectively, with UK/7/91
(the latter strain was isolated from the same chickens as the UK/4/91 strain that was subsequently used
to make an attenuated vaccine; Parsons et al., 1992). The Saudi isolate had 94.7% identity with
UK/7/91. These figures are very similar to the comparison of one of the most recent French isolates
examined, FR/CR94047/94, with UK/7/91 (96.4% identity). Thus the Iranian and Saudi isolates were
clearly of the 793/B type by sequence as well as by serotype.
The Iranian and Saudi isolates had approximately 95% identity in the sequenced region with the 4/91
live vaccinal strain and with FR/CR88061/88, from which the CR88 live vaccine was derived. Thus
the Iranian, Saudi Arabian, French and UK isolates all differed from the two vaccinal strains to a
similar extent: the Iranian and Saudi isolates were not re-isolations of vaccinal virus.
Furthermore our analysis shows that 793/B-serotype strains isolated 15 years apart still had >94%
nucleotide identity in this region.
Host-selected amino acid difference within the S1 protein
Callison et al. (2001) sequenced the S1 gene of the 793/B serotype field isolate UK/4/91 (called
UK/4f/91 in this paper) and its embryo-passaged, attenuated derivative (UK/4e/91). They reported
nucleotide differences at three positions: 283, 1522 and 1589 (where position 1 is the start of the open
reading frame, i.e. inclusive of the signal sequence that is not present in the mature spike protein). We
were unable to confirm the differences at positions 1522 and 1589. At position 283, Callison et al.
(2001) reported a T residue in the field strain and a G residue in the egg-passaged virus. Our
sequencing of PCR products derived from UK/4e/91 reproducibly revealed both G and T at nucleotide
position 283 (Table 1), suggesting that there were two subpopulations in the embryo-passaged virus
with respect to this position - one a T residue as in the initial field virus, the other with a G residue as
7
detected by Callison et al. (2001). Our finding suggested that growth in chickens or embryos favoured
a T or G at nucleotide 283, respectively.
Table 1. Host-dependent variation at nucleotide position 283 (encoding amino acid residue 95) of
the S1 part of the S gene of UK/4/91
Nucleotide at position 283 (amino acid encoded)
Field isolate
UK/4fa/91
T (serine)
Egg-passaged
derivative
UK/4eb/91
T (serine)f
Back passage in chickens
(BP/C) of UK/4e/91
BP/Cc1d
T (serine)f
Passage in embryonated eggs
of BP/C10
BP/C10d
BP/C10/Ee10a
T (serine)
G (alanine)f
G (alanine)f
a
f, field strain of isolate 4/91.
b
e, egg-passage of strain 4/91.
c
C, chicken.
d
Passage number (1 or 10).
e
E, embryonated egg.
f
Sequencing revealed a mixed population comprising both T and G residues.
G (alanine)
The egg-adapted virus had been back-passaged in chickens to establish whether the attenuated
phenotype was stable (i.e. still attenuated); it was. Notwithstanding, it was still possible that backpassage in chickens would favour UK/4/91 virus with a T residue at 283, like the wild-type strain,
even though that was not related to pathogenicity. We therefore sequenced this part of the S gene for
back-passages 1 (BP/C1) and 10 (BP/C10) in chickens of UK/4e/91. BP/C1 still had a mixture of T
and G at this position but in BP/C10 only a T residue was detected. This indicated that either UK/4e/91
had reverted, during passage in chickens, to a T residue (as in the UK/4f/91 field isolate; Table 1), or
that a subpopulation within UK/4e/91 with a T residue had been selected for.
The nucleotides T and G were the first bases in codon 95, encoding serine (field virus, UK/4f/91) and
alanine (embryo-adapted virus, UK/4e/91), respectively. This suggested that growth in chickens and
growth in embryonated eggs was favoured by serine and alanine, respectively, at this position. When
BP/C10 was back-passaged in embryonated eggs, to give BP/C10/E10, position 283 reverted again, to
G, the same as the initial embryo-passaged virus (Table 1). Codon 95 again encoded alanine,
supporting the proposition that UK/4/91 with serine and alanine at this position was somehow better
fitted to replication in chickens and in embryonated eggs, respectively.
Re-inspection of 10 French and 10 UK 793/B-type isolates for which we had sequence data in this
region showed that 19 of them had a T residue at position 283, encoding serine, indicating this to be a
characteristic of field virus, replicating in chickens, although one UK isolate had a leucine residue at
this position.
8
Discussion
Our first evaluation of the degree of variation exhibited within the spike protein genes of a single
serotype was with eight strains of the Massachusetts serotype, collected over a three-decade period
(Cavanagh et al., 1988). The maximum difference between them was 4% of nucleotides and 6% of
amino acids of the S1 protein. Our analysis of S1 of seven isolates of the D274 group, from The
Netherlands and the UK over an 8-year period (Cavanagh et al., 1992) revealed maximum differences
within the group of 3% of nucleotides and 5% of amino acids. In our current investigation of the 793/B
serotype there were maximum differences within the serotype of 4% of nucleotides and 8% of amino
acids. The ratio ds/dns of synonymous to non-synonymous (resulting in amino acid changes) nucleotide
changes for the whole S1 sequences of the 793/B serotype was 1.7, compared with ratios of
approximately 4.4 when Massachusetts, D274 and 793/B serotypes were compared with each other.
For comparison, the ds/dns ratio for the IBV nucleocapsid protein, not expected to be under selective
pressure to change, was 6.5 (Adzhar et al., 1997). ds/dns<1.0 is often cited as evidence for the presence
of positive Darwinian selection. Clearly all the ratios already stated were >1.0, suggesting that some of
the changes were not related to selection pressures. That the ds/dns ratio within the 793/B serotype (1.7)
was nearer to 1.0 than that between other serotypes (mean, 4.4) might indicate that when a serotype
starts to diversify, a high proportion of the small number of S1 mutations are non-synonymous,
whereas subsequent genetic changes involve predominantly synonymous changes.
Interestingly, the seven/eight isolates in the D274 group were not all of the same serotype, as defined
by the virus neutralization test (Cavanagh et al., 1992). Our conclusion then, as now, was that most of
the neutralizing antibody was induced by a small number of dominant epitopes, a small number of
amino acid changes in these regions being sufficient to change the serotype. Serotypes within the D274
group differed by as few as 10 residues in the S1 protein. It may even be that not all of these few
changes were required for the change in serotype.
Do a small number of amino acid changes in S1, sufficient to change the serotype, affect crossprotection? This may sometimes be the case. Isolates UK/123/82 and UK/6/82, of different serotype,
differ by only 3% of S1 amino acids (Cavanagh et al., 1992). When chickens were inoculated with
UK/6/82 and later challenged with UK/123/82, protection against the latter was poor (Cavanagh et al.,
1997). The S1 proteins of the Beaudette and M41 strains of IBV have 95% amino acid identity. When
chickens were inoculated with a recombinant IBV Beaudette expressing the spike protein of the M41
strain, good protection was induced against challenge with M41, as assessed by ciliary activity and
snicking (Hodgson et al., 2004). In contrast, inoculation with Beaudette induced very poor protection
against challenge with M41, suggesting that some of the few amino acids that differed between the two
strains were associated with protection-inducing epitopes.
Some isolates that had 97% S1 nucleotide identity and 95% S1 amino acid identity with the Dutch
D274 strain were assessed as being of different serotypes by neutralization tests using tracheal organ
cultures (Cook, 1984; Cook & Huggins, 1986; Cavanagh et al., 1992). It may be that a given serotype
is always on the brink of becoming a new serotype, as only a small number of changes are required to
bring this about. This is not in conflict with the observation that serotypes exhibit only little variation
in S1 (up to 6% or so) over long periods of time; a sequence investigation of isolates limited to a
particular serotype has, by definition, eliminated isolates that may differ a little more in protein
sequence and that might have, consequently, become a different serotype.
Comparison of the amino acid differences among the UK and French isolates of the 793/B serotype
has not revealed any fixation of changed amino acids (cumulative changes) as time progressed. Also,
some isolates obtained several years apart were more similar than some of those isolated within a
given year. In contrast, Lee & Jackwood (2001) have shown cumulative changes in their study of
9
isolates related to the Delaware serotype. A contributory factor to the different findings might be that
the 793/B isolates in our study had been obtained before the introduction of 793/B-type vaccines,
whereas the Delaware type was in use before and during the period when Lee and Jackwood collected
their isolates.
The change that we observed at residue 95 in the S1 protein of the UK/4/91 strain during passages and
back-passages in embryonated eggs and chickens was not associated with pathogenicity; the attenuated
UK/4e/91 remained non-pathogenic after reversion from an alanine to serine residue had occurred at
position 95. There were two possible explanations for our observation of a switch from a T residue in
the field virus to a G residue in the egg-passaged virus at nucleotide position 283. First, that some of
the field strain had mutated at this position during passage in embryonated eggs. Second, the field
strain itself was a mixture at this position; the ratio of G to T being low enough to make the G residue
not detectable by sequencing of the RT-PCR product. Notwithstanding, replication in chickens and
embryonated eggs was favoured by a serine or alanine residue, respectively, at amino acid position 95.
This situation is reminiscent of that for human influenza virus, in that passage of clinical isolates in
embryonated chicken eggs results in the selection of variants with one, sometimes more, amino acid
substitutions in the haemagglutinin spike protein (Katz & Webster, 1992). The substitutions are in the
vicinity of the receptor binding site. Virus from clinical specimens has an affinity for cell surface
glycoproteins and/or gangliosides with terminal sialyloligosaccharides in which the sialic acid residue
is linked to galactose by an 2,6 linkage. In contrast, influenza virus passaged in embryonated eggs has
an affinity for sialoligosaccharides with terminal 2,3 linkages. Ito et al. (1997) have shown that sialic
acid residues of endodermal cells of the chorioallantoic membrane (CAM) of chicken eggs have 2,3
linkages. The authors concluded that a lack of 2,6 linkages in the CAM of embryonated eggs selected
influenza virus mutants with an haemagglutinin that had a high affinity for 2,3 linkages.
The switching of serine to alanine at residue 95 of the IBV spike protein following passage in
embryonated eggs may or may not be related to interaction with receptors in the CAM of embryonated
eggs. Schultze et al. (1992) showed that attachment of embryo-grown IBV to, and subsequent
agglutination of, chicken erythrocytes required sialic acid that was 2,3-linked to galactose - the
linkage present on CAM cells (Ito et al., 1997). Thus it is conceivable that egg-grown IBV attaches
preferentially to permissive cells that have sialoligosaccharides with terminal 2,3 linkages and that
replication in chickens might favour the selection of spike protein with a higher affinity for 2,6
linkages.
Acknowledgements
This work was supported by the British Chicken Association/British Poultry Council, the Department
of the Environment, Food and Rural Affairs (grant OD0712), Intervet UK, and the Biotechnology and
Biological Sciences Research Council, UK.
10
References

















1. Adzhar, A., Shaw, K., Britton, P. and Cavanagh, D. ((1996)) Universal oligonucleotides for
the detection of infectious bronchitis virus by the polymerase chain reaction. Avian Pathology
25 , pp. 817-836.
2. Adzhar, A., Gough, R. E., Haydon, D., Shaw, K., Britton, P. and Cavanagh, D. ((1997))
Molecular analysis of the 793/B serotype of infectious bronchitis virus in Great Britain. Avian
Pathology 26 , pp. 625-640.
3. Callison, S. A., Jackwood, M. W. and Hilt, D. A. ((2001)) Molecular characterization of
infectious bronchitis virus isolates foreign to the United States and comparison with United
States isolates. Avian Diseases 45 , pp. 492-499. [ crossref ] [ csa ] [ pubmed ]
4. Capua, I., Minta, Z., Karpinska, E., Mawditt, K., Britton, P., Cavanagh, D. and Gough, R.
E. ((1999)) Co-circulation of four types of infectious bronchitis virus (793/B, 624/I, B1648 and
Massachusetts). Avian Pathology 28 , pp. 587-592. [informaworld]
5. Cavanagh, D. ((1997)) Nidovirales: a new order comprising Coronaviridae and Arteriviridae.
Archives of Virology 142 , pp. 629-633. [ csa ] [ pubmed ]
6. Cavanagh, D. ((2001a)) Commentary. A nomenclature for avian coronavirus isolates and the
question of species status. Avian Pathology 30 , pp. 109-115. [informaworld]
7. Cavanagh, D. ((2001b)) Innovation and discovery: the application of nucleic acid-based
technology to avian virus detection and characterization. Avian Pathology 30 , pp. 581-598.
[informaworld]
8. Cavanagh, D. and Naqi, S. ((2003)) Infectious bronchitis. Diseases of Poultry; Iowa State
University Press pp. (pp. 101-119). Ames - In Y.M. Saif, H.J. Barnes, J.R. Glisson, A.M.
Fadly, L.R. McDougald & D.E. Swayne (Eds.); 11th edn
9. Cavanagh, D., Davis, P. J. and Mockett, A. P. A. ((1988)) Amino acids within hypervariable
region 1 of avian coronavirus IBV (Massachusetts serotype) spike glycoprotein are associated
with neutralization epitopes. Virus Research 11 , pp. 141-150. [ crossref ] [ csa ] [ pubmed ]
10. Cavanagh, D., Davis, P. J., Cook, J. K. A., Li, D., Kant, A. and Koch, G. ((1992))
Location of the amino-acid differences in the S1 spike glycoprotein subunit of closely related
serotypes of infectious-bronchitis virus. Avian Pathology 21 , pp. 33-43.
11. Cavanagh, D., Ellis, M. M. and Cook, J. K. A. ((1997)) Relationship between sequence
variation in the S1 spike protein of infectious bronchitis virus and the extent of cross-protection
in vivo. Avian Pathology 26 , pp. 63-74.
12. Cavanagh, D., Mawditt, K., Britton, P. and Naylor, C. J. ((1999)) Longitudinal field
studies of infectious bronchitis virus and avian pneumovirus in broilers using type-specific
polymerase chain reactions. Avian Pathology 28 , pp. 593-605. [informaworld]
13. Cavanagh, D., Mawditt, K., Sharma, M., Drury, S. E., Ainsworth, H. L., Britton, P. and
Gough, R. E. ((2001)) Detection of a coronavirus from turkey poults in Europe genetically
related to infectious bronchitis virus of chickens. Avian Pathology 30 , pp. 355-368.
[informaworld]
14. Cavanagh, D., Mawditt, K., Welchman, D., de, B., Britton, P. and Gough, R. E. ((2002))
Coronaviruses from pheasants (Phasianus colchicus) are genetically closely related to
coronaviruses of domestic fowl (infectious bronchitis virus) and turkeys. Avian Pathology 31 ,
pp. 81-93. [informaworld]
15. Cook, J. K. A. ((1984)) The classification of new serotypes of infectious bronchitis virus
isolated from poultry flocks in Britain between 1981 and 1983. Avian Pathology 13 , pp. 733741.
16. Cook, J. K. A. and Huggins, M. B. ((1986)) Newly isolated serotypes of infectious
bronchitis virus: their role in disease. Avian Pathology 15 , pp. 129-138.
17. Cook, J. K. A., Orbell, S. J., Woods, M. A. and Huggins, M. B. ((1996)) A survey of the
presence of a new infectious bronchitis virus designated 4/91 (793B). Veterinary Record 138 ,
pp. 178-180. [ pubmed ]
11
















18. Cook, J. K. A., Huggins, M. B., Orbell, S. J., Mawditt, K. and Cavanagh, D. ((2001a))
Infectious bronchitis virus vaccine interferes with the replication of avian pneumovirus vaccine
in domestic fowl. Avian Pathology 30 , pp. 233-242. [informaworld]
19. Cook, J. K. A., Chesher, J., Baxendale, W., Greenwood, N., Huggins, M. B. and Orbell,
S. J. ((2001b)) Protection of chickens against renal damage caused by a nephropathogenic
infectious bronchitis virus. Avian Pathology 30 , pp. 423-426. [informaworld]
20. Enjuanes, L., Brian, D., Cavanagh, D., Holmes, K., Lai, M. M. C., Laude, H., Masters,
P., Rottier, P., Siddell, S., Spaan, W. J. M., Taguchi, F. and Talbot, P. ((2000))
Coronaviridae. Virus Taxonomy, Seventh Report of the International Committee on Taxonomy
of Viruses; Academic Press pp. (pp. 835-849). New York - In M.H.V. Van Regenmortel, C.M.
Fauquet, D.H.L. Bishop, E.B. Carstens, M.K. Estes, S.M. Lemon, J. Maniloff, M.A. Mayo, D.J.
McGeoch, C.R. Pringle & R.B. Wickner (Eds.)
21. Farsang, A., Ros, C., Renström, L. H. M., Baule, C., Soós, T. and Belák, S. ((2002))
Molecular epizootiology of infectious bronchitis virus in Sweden indicating the involvement of
a vaccine strain. Avian Pathology 31 , pp. 229-236. [informaworld]
22. Felsentein, J. ((1993)) PHYLIP (Phylogeny Inference Package); Department of Genetics,
University of Washington SeattleWA - version 3.5c
23. Gough, R. E., Randall, C. J., Dagless, M., Alexander, D. J., Cox, W. J. and Pearson, D.
((1992)) A 'new' strain of infectious bronchitis virus infecting domestic fowl in Great Britain.
The Veterinary Record 130
24. Guy, J. S. ((2000)) Turkey coronavirus is more closely related to avian infectious bronchitis
virus than to mammalian coronaviruses. Avian Pathology 29 , pp. 206-212. [informaworld]
25. Hodgson, T., Casai, R., Dove, B., Britton, P. and Cavanagh, D. ((2004)) Recombinant
infectious bronchitis coronavirus Beaudette with the spike protein gene of the pathogenic M41
strain remains attenuated but induces protective immunity. Journal of Virology 78 , pp. 1380413811. [ csa ] [ pubmed ] [ crossref ]
26. Ito, T., Suzuki, Y., Takada, A., Kawamoto, A., Otsuki, K., Masuda, H., Yamada, M.,
Suzuki, T., Kida, H. and Kawaoka, Y. ((1997)) Differences in sialic acid-galactose linkages in
the chicken amnion and allantois influence human influenza virus receptor specificity and
variant selection. Journal of Virology 71 , pp. 3357-3362. [ csa ] [ pubmed ]
27. Kant, A., Koch, G., van Roozelaar, D. J., Kusters, J. G., Poelwijk, F. A. J. and van der
Zeijst, B. A. M. ((1992)) Location of antigenic sites defined by neutralizing monclonal
antibodies on the S1 avian infectious bronchitis virus glycopolypeptide. Journal of General
Virology 73 , pp. 591-596. [ pubmed ]
28. Katz, J. M. and Webster, R. G. ((1992)) Amino acid sequence identity between the HA1 of
influenza A (H3N2) viruses grown in mammalian and primary chick kidney cells. Journal of
General Virology 73 , pp. 1159-1165. [ pubmed ] [ csa ]
29. Koch, G., Hartog, L., Kant, A. and van Roozelaar, D. J. ((1990)) Antigenic domains on
the peplomer protein of avian infectious bronchitis virus: correlation with biological functions.
Journal of General Virology 71 , pp. 1929-1935. [ pubmed ] [ csa ]
30. Lee, C. W. and Jackwood, M. W. ((2001)) Origin and evolution of Georgia 98 (GA98), a
new serotype of avian infectious bronchitis virus. Virus Research 80 , pp. 33-39. [ csa ] [
pubmed ] [ crossref ]
31. Li, H. and Yang, H-C. ((2001)) Sequence analysis of nephropathogenic infectious
bronchitis virus strains of the Massachusetts genotype in Beijing. Avian Pathology 30 , pp.
535-541. [informaworld]
32. Matthijs, M. R. G., van Eck, J. H. H., Landman, W. J. M. and Stegeman, J. A. ((2003))
Ability of Massachusetts-type infectious bronchitis virus to increase colibacillosis
susceptibility in commercial broilers: a comparison between vaccine and virulent field virus.
Avian Pathology 32 , pp. 473-481. [ pubmed ] [ crossref ]
33. Meulemans, G., Boschmans, M., Decaesstecker, M., Berg, T.P. van den, Denis, P. and
Cavanagh, D. ((2001)) Epidemiology of infectious bronchitis virus in Belgian broilers: a
retrospective study, 1986 to 1995. Avian Pathology 30 , pp. 411-421. [informaworld]
12





34. Parsons, D., Ellis, M. M., Cavanagh, D. and Cook, J. K. A. ((1992)) Characterisation of an
avian infectious bronchitis virus isolated from IB-vaccinated broiler breeder flocks. The
Veterinary Record 131 , pp. 408-411. [ pubmed ]
35. Picault, J. P., Drouin, P., Lamande, J., Allee, C., Toux, J. Y., Coq, H. Le, Guittet, M.
and Bennejean, G. ((1995); 28-29, March) L'epizootie recente de bronchite infectieuse aviaire
en France: importance, evolution et etiologie. Proceedings of the 1eres Journée de la
Recherche Avicole, Angers pp. 177-179.
36. Schultze, B., Cavanagh, D. and Herrler, G. ((1992)) Neuraminidase treatment of avian
infectious bronchitis coronavirus reveals a hemagglutinating activity that is dependent on sialic
acid-containing receptors on erythrocytes. Virology 189 , pp. 792-794. [ csa ] [ pubmed ] [
crossref ]
37. Thompson, J. D., Gibson, T. J., Plewniak, F., Jeanmougin, F. and Higgins, D. G. ((1994))
The clustal_X windows interface: flexible strategies for multiple sequence alignment aided by
quality analysis tools. Nucleic Acids Research 25 , pp. 4876-4882. [ crossref ]
38. Welchman, D., Bradbury, J. M., Cavanagh, D. and Aebischer, N. J. ((2002)) Infectious
agents associated with respiratory disease in pheasants. Veterinary Record 150 , pp. 658-664. [
pubmed ]
-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------