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Comparative features of infections of two Massachusetts (Mass) infectious bronchitis virus (IBV) variants isolated from Western Canadian layer flocks

Identifieur interne : 000224 ( Pmc/Corpus ); précédent : 000223; suivant : 000225

Comparative features of infections of two Massachusetts (Mass) infectious bronchitis virus (IBV) variants isolated from Western Canadian layer flocks

Auteurs : Aruna Amarasinghe ; Upasama De Silva Senapathi ; Mohamed Sarjoon Abdul-Cader ; Shelly Popowich ; Frank Marshall ; Susan C. Cork ; Frank Van Der Meer ; Susantha Gomis ; Mohamed Faizal Abdul-Careem

Source :

RBID : PMC:6288874

Abstract

Background

Infectious bronchitis virus (IBV) is one of the leading causes of mortality and morbidity in chickens. There are numerous serotypes and variants, which do not confer cross protection resulting in failure of currently used IBV vaccines. Although variant IBV isolates with major genetic differences have been subjected to comparative studies, it is unknown whether minor genetic differences in IBV variants within a serotype are different in terms of pathogenesis and eliciting host responses. Two Massachusetts (Mass) variant IBV isolates recovered from commercial layer flocks in the Western Canadian provinces of Alberta (AB) and Saskatchewan (SK) were compared genetically and evaluated for their pathogenicity, tissue distribution and ability to recruit and replicate in macrophages.

Results

Although whole genome sequencing of these two Mass IBV isolates showed low similarity with the M41 vaccinal strain, they had an identical nucleotide sequence at open reading frames (ORFs) 3a, 3b, envelop (E), matrix (M), 5a and 5b. The rest of the ORFs of these 2 IBV isolates showed 99.9% nucleotide similarity. However, upon experimental infection, we found that the IBV isolate originating from AB was different to the one that originated in SK due to higher tracheal lesion scores and lower lung viral replication and lower genome loads in cecal tonsils. Nevertheless, both IBV isolates elicited host responses characterized by significant macrophage recruitment to the respiratory tract and there was evidence that both IBV isolates replicated within tracheal and lung macrophages.

Conclusions

Overall, this study shows that Mass variant IBV isolates, although possessing minor genetic variations, can lead to significant differences in pathogenicity in young chickens. Further studies are required to investigate the pathogenicity of these two Mass variant IBV isolates in laying hens.

Electronic supplementary material

The online version of this article (10.1186/s12917-018-1720-9) contains supplementary material, which is available to authorized users.


Url:
DOI: 10.1186/s12917-018-1720-9
PubMed: 30526618
PubMed Central: 6288874

Links to Exploration step

PMC:6288874

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<name sortKey="Abdul Careem, Mohamed Faizal" sort="Abdul Careem, Mohamed Faizal" uniqKey="Abdul Careem M" first="Mohamed Faizal" last="Abdul-Careem">Mohamed Faizal Abdul-Careem</name>
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<title>Background</title>
<p id="Par1">Infectious bronchitis virus (IBV) is one of the leading causes of mortality and morbidity in chickens. There are numerous serotypes and variants, which do not confer cross protection resulting in failure of currently used IBV vaccines. Although variant IBV isolates with major genetic differences have been subjected to comparative studies, it is unknown whether minor genetic differences in IBV variants within a serotype are different in terms of pathogenesis and eliciting host responses. Two Massachusetts (Mass) variant IBV isolates recovered from commercial layer flocks in the Western Canadian provinces of Alberta (AB) and Saskatchewan (SK) were compared genetically and evaluated for their pathogenicity, tissue distribution and ability to recruit and replicate in macrophages.</p>
</sec>
<sec>
<title>Results</title>
<p id="Par2">Although whole genome sequencing of these two Mass IBV isolates showed low similarity with the M41 vaccinal strain, they had an identical nucleotide sequence at open reading frames (ORFs) 3a, 3b, envelop (E), matrix (M), 5a and 5b. The rest of the ORFs of these 2 IBV isolates showed 99.9% nucleotide similarity. However, upon experimental infection, we found that the IBV isolate originating from AB was different to the one that originated in SK due to higher tracheal lesion scores and lower lung viral replication and lower genome loads in cecal tonsils. Nevertheless, both IBV isolates elicited host responses characterized by significant macrophage recruitment to the respiratory tract and there was evidence that both IBV isolates replicated within tracheal and lung macrophages.</p>
</sec>
<sec>
<title>Conclusions</title>
<p id="Par3">Overall, this study shows that Mass variant IBV isolates, although possessing minor genetic variations, can lead to significant differences in pathogenicity in young chickens. Further studies are required to investigate the pathogenicity of these two Mass variant IBV isolates in laying hens.</p>
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<sec>
<title>Electronic supplementary material</title>
<p>The online version of this article (10.1186/s12917-018-1720-9) contains supplementary material, which is available to authorized users.</p>
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<listBibl>
<biblStruct>
<analytic>
<author>
<name sortKey="Cavanagh, D" uniqKey="Cavanagh D">D Cavanagh</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Cavanagh, D" uniqKey="Cavanagh D">D Cavanagh</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Cumming, Rb" uniqKey="Cumming R">RB Cumming</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Kingham, Bf" uniqKey="Kingham B">BF Kingham</name>
</author>
<author>
<name sortKey="Keeler, Cl" uniqKey="Keeler C">CL Keeler</name>
</author>
<author>
<name sortKey="Nix, Wa" uniqKey="Nix W">WA Nix</name>
</author>
<author>
<name sortKey="Ladman, Bs" uniqKey="Ladman B">BS Ladman</name>
</author>
<author>
<name sortKey="Gelb, J" uniqKey="Gelb J">J Gelb</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Song, C S" uniqKey="Song C">C-S Song</name>
</author>
<author>
<name sortKey="Lee, Y J" uniqKey="Lee Y">Y-J Lee</name>
</author>
<author>
<name sortKey="Lee, C W" uniqKey="Lee C">C-W Lee</name>
</author>
<author>
<name sortKey="Sung, H W" uniqKey="Sung H">H-W Sung</name>
</author>
<author>
<name sortKey="Kim, J H" uniqKey="Kim J">J-H Kim</name>
</author>
<author>
<name sortKey="Mo, I P" uniqKey="Mo I">I-P Mo</name>
</author>
<author>
<name sortKey="Izumiya, Y" uniqKey="Izumiya Y">Y Izumiya</name>
</author>
<author>
<name sortKey="Jang, H K" uniqKey="Jang H">H-K Jang</name>
</author>
<author>
<name sortKey="Mikami, T" uniqKey="Mikami T">T Mikami</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Liu, S" uniqKey="Liu S">S Liu</name>
</author>
<author>
<name sortKey="Chen, J" uniqKey="Chen J">J Chen</name>
</author>
<author>
<name sortKey="Han, Z" uniqKey="Han Z">Z Han</name>
</author>
<author>
<name sortKey="Zhang, Q" uniqKey="Zhang Q">Q Zhang</name>
</author>
<author>
<name sortKey="Shao, Y" uniqKey="Shao Y">Y Shao</name>
</author>
<author>
<name sortKey="Kong, X" uniqKey="Kong X">X Kong</name>
</author>
<author>
<name sortKey="Tong, G" uniqKey="Tong G">G Tong</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Reddy, Vrap" uniqKey="Reddy V">VRAP Reddy</name>
</author>
<author>
<name sortKey="Theuns, S" uniqKey="Theuns S">S Theuns</name>
</author>
<author>
<name sortKey="Roukaerts, Idm" uniqKey="Roukaerts I">IDM Roukaerts</name>
</author>
<author>
<name sortKey="Zeller, M" uniqKey="Zeller M">M Zeller</name>
</author>
<author>
<name sortKey="Matthijnssens, J" uniqKey="Matthijnssens J">J Matthijnssens</name>
</author>
<author>
<name sortKey="Nauwynck, Hj" uniqKey="Nauwynck H">HJ Nauwynck</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Grgic, H" uniqKey="Grgic H">H Grgic</name>
</author>
<author>
<name sortKey="Hunter, Db" uniqKey="Hunter D">DB Hunter</name>
</author>
<author>
<name sortKey="Hunton, P" uniqKey="Hunton P">P Hunton</name>
</author>
<author>
<name sortKey="Nagy, E" uniqKey="Nagy E">E Nagy</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Boroomand, Z" uniqKey="Boroomand Z">Z Boroomand</name>
</author>
<author>
<name sortKey="Asasi, K" uniqKey="Asasi K">K Asasi</name>
</author>
<author>
<name sortKey="Mohammadi, A" uniqKey="Mohammadi A">A Mohammadi</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Franca, M" uniqKey="Franca M">M Franca</name>
</author>
<author>
<name sortKey="Woolcock, Pr" uniqKey="Woolcock P">PR Woolcock</name>
</author>
<author>
<name sortKey="Yu, M" uniqKey="Yu M">M Yu</name>
</author>
<author>
<name sortKey="Jackwood, Mw" uniqKey="Jackwood M">MW Jackwood</name>
</author>
<author>
<name sortKey="Shivaprasad, Hl" uniqKey="Shivaprasad H">HL Shivaprasad</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Ambali, Ag" uniqKey="Ambali A">AG Ambali</name>
</author>
<author>
<name sortKey="Jones, Rc" uniqKey="Jones R">RC Jones</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="El Houadfi, M" uniqKey="El Houadfi M">M El-Houadfi</name>
</author>
<author>
<name sortKey="Jones, Rc" uniqKey="Jones R">RC Jones</name>
</author>
<author>
<name sortKey="Cook, Jka" uniqKey="Cook J">JKA Cook</name>
</author>
<author>
<name sortKey="Ambali, Ag" uniqKey="Ambali A">AG Ambali</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Sevoian, M" uniqKey="Sevoian M">M Sevoian</name>
</author>
<author>
<name sortKey="Levine, P" uniqKey="Levine P">P Levine</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Macdonald, Jw" uniqKey="Macdonald J">JW MacDonald</name>
</author>
<author>
<name sortKey="Mcmartin, Da" uniqKey="Mcmartin D">DA McMartin</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Raj, Gd" uniqKey="Raj G">GD Raj</name>
</author>
<author>
<name sortKey="Jones, Rc" uniqKey="Jones R">RC Jones</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Cook, Jk" uniqKey="Cook J">JK Cook</name>
</author>
<author>
<name sortKey="Jackwood, M" uniqKey="Jackwood M">M Jackwood</name>
</author>
<author>
<name sortKey="Jones, Rc" uniqKey="Jones R">RC Jones</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Crinion, Rap" uniqKey="Crinion R">RAP Crinion</name>
</author>
<author>
<name sortKey="Ball, Ra" uniqKey="Ball R">RA Ball</name>
</author>
<author>
<name sortKey="Hofstad, Ms" uniqKey="Hofstad M">MS Hofstad</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Benyeda, Z" uniqKey="Benyeda Z">Z Benyeda</name>
</author>
<author>
<name sortKey="Mato, T" uniqKey="Mato T">T Mato</name>
</author>
<author>
<name sortKey="Suveges, T" uniqKey="Suveges T">T Suveges</name>
</author>
<author>
<name sortKey="Szabo, E" uniqKey="Szabo E">E Szabo</name>
</author>
<author>
<name sortKey="Kardi, V" uniqKey="Kardi V">V Kardi</name>
</author>
<author>
<name sortKey="Abonyi Toth, Z" uniqKey="Abonyi Toth Z">Z Abonyi-Toth</name>
</author>
<author>
<name sortKey="Rusvai, M" uniqKey="Rusvai M">M Rusvai</name>
</author>
<author>
<name sortKey="Palya, V" uniqKey="Palya V">V Palya</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Boltz, Da" uniqKey="Boltz D">DA Boltz</name>
</author>
<author>
<name sortKey="Nakai, M" uniqKey="Nakai M">M Nakai</name>
</author>
<author>
<name sortKey="Bahra, Jm" uniqKey="Bahra J">JM Bahra</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Liu, S" uniqKey="Liu S">S Liu</name>
</author>
<author>
<name sortKey="Zhang, X" uniqKey="Zhang X">X Zhang</name>
</author>
<author>
<name sortKey="Wang, Y" uniqKey="Wang Y">Y Wang</name>
</author>
<author>
<name sortKey="Li, C" uniqKey="Li C">C Li</name>
</author>
<author>
<name sortKey="Han, Z" uniqKey="Han Z">Z Han</name>
</author>
<author>
<name sortKey="Shao, Y" uniqKey="Shao Y">Y Shao</name>
</author>
<author>
<name sortKey="Li, H" uniqKey="Li H">H Li</name>
</author>
<author>
<name sortKey="Kong, X" uniqKey="Kong X">X Kong</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Alvarado, Ir" uniqKey="Alvarado I">IR Alvarado</name>
</author>
<author>
<name sortKey="Villegas, P" uniqKey="Villegas P">P Villegas</name>
</author>
<author>
<name sortKey="Mossos, N" uniqKey="Mossos N">N Mossos</name>
</author>
<author>
<name sortKey="Jackwood, Mw" uniqKey="Jackwood M">MW Jackwood</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Kusters, Jg" uniqKey="Kusters J">JG Kusters</name>
</author>
<author>
<name sortKey="Niesters, Hgm" uniqKey="Niesters H">HGM Niesters</name>
</author>
<author>
<name sortKey="Bleumink Pluym, Nmc" uniqKey="Bleumink Pluym N">NMC Bleumink-Pluym</name>
</author>
<author>
<name sortKey="Davelaar, Fg" uniqKey="Davelaar F">FG Davelaar</name>
</author>
<author>
<name sortKey="Horzinek, Mc" uniqKey="Horzinek M">MC Horzinek</name>
</author>
<author>
<name sortKey="Van Der Zeijst, Bam" uniqKey="Van Der Zeijst B">BAM Van der Zeijst</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Lee, C W" uniqKey="Lee C">C-W Lee</name>
</author>
<author>
<name sortKey="Jackwood, Mw" uniqKey="Jackwood M">MW Jackwood</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Kottier, Sa" uniqKey="Kottier S">SA Kottier</name>
</author>
<author>
<name sortKey="Cavanagh, D" uniqKey="Cavanagh D">D Cavanagh</name>
</author>
<author>
<name sortKey="Britton, P" uniqKey="Britton P">P Britton</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Wang, L" uniqKey="Wang L">L Wang</name>
</author>
<author>
<name sortKey="Xu, Y" uniqKey="Xu Y">Y Xu</name>
</author>
<author>
<name sortKey="Collisson, Ew" uniqKey="Collisson E">EW Collisson</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Mardani, K" uniqKey="Mardani K">K Mardani</name>
</author>
<author>
<name sortKey="Noormohammadi, A" uniqKey="Noormohammadi A">A Noormohammadi</name>
</author>
<author>
<name sortKey="Ignjatovic, J" uniqKey="Ignjatovic J">J Ignjatovic</name>
</author>
<author>
<name sortKey="Browning, G" uniqKey="Browning G">G Browning</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Jia, W" uniqKey="Jia W">W Jia</name>
</author>
<author>
<name sortKey="Karaca, K" uniqKey="Karaca K">K Karaca</name>
</author>
<author>
<name sortKey="Parrish, Cr" uniqKey="Parrish C">CR Parrish</name>
</author>
<author>
<name sortKey="Naqi, Sa" uniqKey="Naqi S">SA Naqi</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Montassier, Hj" uniqKey="Montassier H">HJ Montassier</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Cavanagh, D" uniqKey="Cavanagh D">D Cavanagh</name>
</author>
<author>
<name sortKey="Davis, Pj" uniqKey="Davis P">PJ Davis</name>
</author>
<author>
<name sortKey="Darbyshire, Jh" uniqKey="Darbyshire J">JH Darbyshire</name>
</author>
<author>
<name sortKey="Peters, Rw" uniqKey="Peters R">RW Peters</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Cavanagh, D" uniqKey="Cavanagh D">D Cavanagh</name>
</author>
<author>
<name sortKey="Davis, Pj" uniqKey="Davis P">PJ Davis</name>
</author>
<author>
<name sortKey="Cook, Jka" uniqKey="Cook J">JKA Cook</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Selim, K" uniqKey="Selim K">K Selim</name>
</author>
<author>
<name sortKey="Arafa, As" uniqKey="Arafa A">AS Arafa</name>
</author>
<author>
<name sortKey="Hussein, Ha" uniqKey="Hussein H">HA Hussein</name>
</author>
<author>
<name sortKey="El Sanousi, Aa" uniqKey="El Sanousi A">AA El-Sanousi</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Awad, F" uniqKey="Awad F">F Awad</name>
</author>
<author>
<name sortKey="Baylis, M" uniqKey="Baylis M">M Baylis</name>
</author>
<author>
<name sortKey="Ganapathy, K" uniqKey="Ganapathy K">K Ganapathy</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Martin, Ea" uniqKey="Martin E">EA Martin</name>
</author>
<author>
<name sortKey="Brash, Ml" uniqKey="Brash M">ML Brash</name>
</author>
<author>
<name sortKey="Hoyland, Sk" uniqKey="Hoyland S">SK Hoyland</name>
</author>
<author>
<name sortKey="Coventry, Jm" uniqKey="Coventry J">JM Coventry</name>
</author>
<author>
<name sortKey="Sandrock, C" uniqKey="Sandrock C">C Sandrock</name>
</author>
<author>
<name sortKey="Guerin, Mt" uniqKey="Guerin M">MT Guerin</name>
</author>
<author>
<name sortKey="Ojkic, D" uniqKey="Ojkic D">D Ojkic</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Jackwood, Mw" uniqKey="Jackwood M">MW Jackwood</name>
</author>
<author>
<name sortKey="Lee, Dh" uniqKey="Lee D">DH Lee</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Hong, S M" uniqKey="Hong S">S-M Hong</name>
</author>
<author>
<name sortKey="Kwon, H J" uniqKey="Kwon H">H-J Kwon</name>
</author>
<author>
<name sortKey="Kim, I H" uniqKey="Kim I">I-H Kim</name>
</author>
<author>
<name sortKey="Mo, M L" uniqKey="Mo M">M-L Mo</name>
</author>
<author>
<name sortKey="Kim, J H" uniqKey="Kim J">J-H Kim</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Marandino, A" uniqKey="Marandino A">A Marandino</name>
</author>
<author>
<name sortKey="Pereda, A" uniqKey="Pereda A">A Pereda</name>
</author>
<author>
<name sortKey="Tomas, G" uniqKey="Tomas G">G Tomas</name>
</author>
<author>
<name sortKey="Hernandez, M" uniqKey="Hernandez M">M Hernandez</name>
</author>
<author>
<name sortKey="Iraola, G" uniqKey="Iraola G">G Iraola</name>
</author>
<author>
<name sortKey="Craig, Mi" uniqKey="Craig M">MI Craig</name>
</author>
<author>
<name sortKey="Hernandez, D" uniqKey="Hernandez D">D Hernandez</name>
</author>
<author>
<name sortKey="Banda, A" uniqKey="Banda A">A Banda</name>
</author>
<author>
<name sortKey="Villegas, P" uniqKey="Villegas P">P Villegas</name>
</author>
<author>
<name sortKey="Panzera, Y" uniqKey="Panzera Y">Y Panzera</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Smati, R" uniqKey="Smati R">R Smati</name>
</author>
<author>
<name sortKey="Silim, A" uniqKey="Silim A">A Silim</name>
</author>
<author>
<name sortKey="Guertin, C" uniqKey="Guertin C">C Guertin</name>
</author>
<author>
<name sortKey="Henrichon, M" uniqKey="Henrichon M">M Henrichon</name>
</author>
<author>
<name sortKey="Marandi, M" uniqKey="Marandi M">M Marandi</name>
</author>
<author>
<name sortKey="Arella, M" uniqKey="Arella M">M Arella</name>
</author>
<author>
<name sortKey="Merzouki, A" uniqKey="Merzouki A">A Merzouki</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Feng, K" uniqKey="Feng K">K Feng</name>
</author>
<author>
<name sortKey="Wang, F" uniqKey="Wang F">F Wang</name>
</author>
<author>
<name sortKey="Xue, Y" uniqKey="Xue Y">Y Xue</name>
</author>
<author>
<name sortKey="Zhou, Q" uniqKey="Zhou Q">Q Zhou</name>
</author>
<author>
<name sortKey="Chen, F" uniqKey="Chen F">F Chen</name>
</author>
<author>
<name sortKey="Bi, Y" uniqKey="Bi Y">Y Bi</name>
</author>
<author>
<name sortKey="Xie, Q" uniqKey="Xie Q">Q Xie</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Mo, Ml" uniqKey="Mo M">ML Mo</name>
</author>
<author>
<name sortKey="Li, M" uniqKey="Li M">M Li</name>
</author>
<author>
<name sortKey="Huang, Bc" uniqKey="Huang B">BC Huang</name>
</author>
<author>
<name sortKey="Fan, Ws" uniqKey="Fan W">WS Fan</name>
</author>
<author>
<name sortKey="Wei, P" uniqKey="Wei P">P Wei</name>
</author>
<author>
<name sortKey="Wei, Tc" uniqKey="Wei T">TC Wei</name>
</author>
<author>
<name sortKey="Cheng, Qy" uniqKey="Cheng Q">QY Cheng</name>
</author>
<author>
<name sortKey="Wei, Zj" uniqKey="Wei Z">ZJ Wei</name>
</author>
<author>
<name sortKey="Lang, Yh" uniqKey="Lang Y">YH Lang</name>
</author>
</analytic>
</biblStruct>
<biblStruct></biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Van Beurden, Sj" uniqKey="Van Beurden S">SJ van Beurden</name>
</author>
<author>
<name sortKey="Berends, Aj" uniqKey="Berends A">AJ Berends</name>
</author>
<author>
<name sortKey="Kramer Kuhl, A" uniqKey="Kramer Kuhl A">A Kramer-Kuhl</name>
</author>
<author>
<name sortKey="Spekreijse, D" uniqKey="Spekreijse D">D Spekreijse</name>
</author>
<author>
<name sortKey="Chenard, G" uniqKey="Chenard G">G Chenard</name>
</author>
<author>
<name sortKey="Philipp, Hc" uniqKey="Philipp H">HC Philipp</name>
</author>
<author>
<name sortKey="Mundt, E" uniqKey="Mundt E">E Mundt</name>
</author>
<author>
<name sortKey="Rottier, Pjm" uniqKey="Rottier P">PJM Rottier</name>
</author>
<author>
<name sortKey="Verheije, Mh" uniqKey="Verheije M">MH Verheije</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Benyeda, Z" uniqKey="Benyeda Z">Z Benyeda</name>
</author>
<author>
<name sortKey="Szeredi, L" uniqKey="Szeredi L">L Szeredi</name>
</author>
<author>
<name sortKey="Mat, T" uniqKey="Mat T">T Mató</name>
</author>
<author>
<name sortKey="Suveges, T" uniqKey="Suveges T">T Süveges</name>
</author>
<author>
<name sortKey="Balka, G" uniqKey="Balka G">G Balka</name>
</author>
<author>
<name sortKey="Abonyi T Th, Z" uniqKey="Abonyi T Th Z">Z Abonyi-Tóth</name>
</author>
<author>
<name sortKey="Rusvai, M" uniqKey="Rusvai M">M Rusvai</name>
</author>
<author>
<name sortKey="Palya, V" uniqKey="Palya V">V Palya</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="W Q, F" uniqKey="W Q F">F W-q</name>
</author>
<author>
<name sortKey="H N, W" uniqKey="H N W">W H-n</name>
</author>
<author>
<name sortKey="Zhang, Y" uniqKey="Zhang Y">Y Zhang</name>
</author>
<author>
<name sortKey="Z B, G" uniqKey="Z B G">G Z-b</name>
</author>
<author>
<name sortKey="Wang, T" uniqKey="Wang T">T Wang</name>
</author>
<author>
<name sortKey="C W, X" uniqKey="C W X">X C-w</name>
</author>
<author>
<name sortKey="A Y, Z" uniqKey="A Y Z">Z A-y</name>
</author>
<author>
<name sortKey="Yang, X" uniqKey="Yang X">X Yang</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Cheng, J" uniqKey="Cheng J">J Cheng</name>
</author>
<author>
<name sortKey="Huo, C" uniqKey="Huo C">C Huo</name>
</author>
<author>
<name sortKey="Zhao, J" uniqKey="Zhao J">J Zhao</name>
</author>
<author>
<name sortKey="Liu, T" uniqKey="Liu T">T Liu</name>
</author>
<author>
<name sortKey="Li, X" uniqKey="Li X">X Li</name>
</author>
<author>
<name sortKey="Yan, S" uniqKey="Yan S">S Yan</name>
</author>
<author>
<name sortKey="Wang, Z" uniqKey="Wang Z">Z Wang</name>
</author>
<author>
<name sortKey="Hu, Y" uniqKey="Hu Y">Y Hu</name>
</author>
<author>
<name sortKey="Zhang, G" uniqKey="Zhang G">G Zhang</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Chhabra, R" uniqKey="Chhabra R">R Chhabra</name>
</author>
<author>
<name sortKey="Kuchipudi, Sv" uniqKey="Kuchipudi S">SV Kuchipudi</name>
</author>
<author>
<name sortKey="Chantrey, J" uniqKey="Chantrey J">J Chantrey</name>
</author>
<author>
<name sortKey="Ganapathy, K" uniqKey="Ganapathy K">K Ganapathy</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Chhabra, R" uniqKey="Chhabra R">R Chhabra</name>
</author>
<author>
<name sortKey="Ball, C" uniqKey="Ball C">C Ball</name>
</author>
<author>
<name sortKey="Chantrey, J" uniqKey="Chantrey J">J Chantrey</name>
</author>
<author>
<name sortKey="Ganapathy, K" uniqKey="Ganapathy K">K Ganapathy</name>
</author>
</analytic>
</biblStruct>
<biblStruct></biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Cavanagh, D" uniqKey="Cavanagh D">D Cavanagh</name>
</author>
<author>
<name sortKey="Davis, Pj" uniqKey="Davis P">PJ Davis</name>
</author>
<author>
<name sortKey="Cook, Jk" uniqKey="Cook J">JK Cook</name>
</author>
<author>
<name sortKey="Li, D" uniqKey="Li D">D Li</name>
</author>
<author>
<name sortKey="Kant, A" uniqKey="Kant A">A Kant</name>
</author>
<author>
<name sortKey="Koch, G" uniqKey="Koch G">G Koch</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Binns, Mm" uniqKey="Binns M">MM Binns</name>
</author>
<author>
<name sortKey="Boursnell, Me" uniqKey="Boursnell M">ME Boursnell</name>
</author>
<author>
<name sortKey="Tomley, Fm" uniqKey="Tomley F">FM Tomley</name>
</author>
<author>
<name sortKey="Brown, Dk" uniqKey="Brown D">DK Brown</name>
</author>
</analytic>
</biblStruct>
<biblStruct></biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Stachowiak, Bkd" uniqKey="Stachowiak B">BKD Stachowiak</name>
</author>
<author>
<name sortKey="Hunton, P" uniqKey="Hunton P">P Hunton</name>
</author>
<author>
<name sortKey="Gillingham, S" uniqKey="Gillingham S">S Gillingham</name>
</author>
<author>
<name sortKey="Nagy, E" uniqKey="Nagy E">É Nagy</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Ignjatovic, J" uniqKey="Ignjatovic J">J Ignjatovic</name>
</author>
<author>
<name sortKey="Mcwaters, Pg" uniqKey="Mcwaters P">PG McWaters</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Kant, A" uniqKey="Kant A">A Kant</name>
</author>
<author>
<name sortKey="Koch, G" uniqKey="Koch G">G Koch</name>
</author>
<author>
<name sortKey="Van Roozelaar, Dj" uniqKey="Van Roozelaar D">DJ van Roozelaar</name>
</author>
<author>
<name sortKey="Kusters, Jg" uniqKey="Kusters J">JG Kusters</name>
</author>
<author>
<name sortKey="Poelwijk, Faj" uniqKey="Poelwijk F">FAJ Poelwijk</name>
</author>
<author>
<name sortKey="Van Der Zeijst, Bam" uniqKey="Van Der Zeijst B">BAM van der Zeijst</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Cavanagh, D" uniqKey="Cavanagh D">D Cavanagh</name>
</author>
<author>
<name sortKey="Davis, Pj" uniqKey="Davis P">PJ Davis</name>
</author>
<author>
<name sortKey="Mockett, Apa" uniqKey="Mockett A">APA Mockett</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Quinteros, Ja" uniqKey="Quinteros J">JA Quinteros</name>
</author>
<author>
<name sortKey="Lee, S W" uniqKey="Lee S">S-W Lee</name>
</author>
<author>
<name sortKey="Markham, Pf" uniqKey="Markham P">PF Markham</name>
</author>
<author>
<name sortKey="Noormohammadi, Ah" uniqKey="Noormohammadi A">AH Noormohammadi</name>
</author>
<author>
<name sortKey="Hartley, Ca" uniqKey="Hartley C">CA Hartley</name>
</author>
<author>
<name sortKey="Legione, Ar" uniqKey="Legione A">AR Legione</name>
</author>
<author>
<name sortKey="Coppo, Mjc" uniqKey="Coppo M">MJC Coppo</name>
</author>
<author>
<name sortKey="Vaz, Pk" uniqKey="Vaz P">PK Vaz</name>
</author>
<author>
<name sortKey="Browning, Gf" uniqKey="Browning G">GF Browning</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Abozeid, Hh" uniqKey="Abozeid H">HH Abozeid</name>
</author>
<author>
<name sortKey="Paldurai, A" uniqKey="Paldurai A">A Paldurai</name>
</author>
<author>
<name sortKey="Khattar, Sk" uniqKey="Khattar S">SK Khattar</name>
</author>
<author>
<name sortKey="Afifi, Ma" uniqKey="Afifi M">MA Afifi</name>
</author>
<author>
<name sortKey="El Kady, Mf" uniqKey="El Kady M">MF El-Kady</name>
</author>
<author>
<name sortKey="El Deeb, Ah" uniqKey="El Deeb A">AH El-Deeb</name>
</author>
<author>
<name sortKey="Samal, Sk" uniqKey="Samal S">SK Samal</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Naguib, Mm" uniqKey="Naguib M">MM Naguib</name>
</author>
<author>
<name sortKey="Hoper, D" uniqKey="Hoper D">D Höper</name>
</author>
<author>
<name sortKey="Arafa, A S" uniqKey="Arafa A">A-S Arafa</name>
</author>
<author>
<name sortKey="Setta, Am" uniqKey="Setta A">AM Setta</name>
</author>
<author>
<name sortKey="Abed, M" uniqKey="Abed M">M Abed</name>
</author>
<author>
<name sortKey="Monne, I" uniqKey="Monne I">I Monne</name>
</author>
<author>
<name sortKey="Beer, M" uniqKey="Beer M">M Beer</name>
</author>
<author>
<name sortKey="Harder, Tc" uniqKey="Harder T">TC Harder</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Wu, X" uniqKey="Wu X">X Wu</name>
</author>
<author>
<name sortKey="Yang, X" uniqKey="Yang X">X Yang</name>
</author>
<author>
<name sortKey="Xu, P" uniqKey="Xu P">P Xu</name>
</author>
<author>
<name sortKey="Zhou, L" uniqKey="Zhou L">L Zhou</name>
</author>
<author>
<name sortKey="Zhang, Z" uniqKey="Zhang Z">Z Zhang</name>
</author>
<author>
<name sortKey="Wang, H" uniqKey="Wang H">H Wang</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="X L, L" uniqKey="X L L">L X-l</name>
</author>
<author>
<name sortKey="J L, S" uniqKey="J L S">S J-l</name>
</author>
<author>
<name sortKey="J X, Z" uniqKey="J X Z">Z J-x</name>
</author>
<author>
<name sortKey="G Z, Z" uniqKey="G Z Z">Z G-z</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Khataby, K" uniqKey="Khataby K">K Khataby</name>
</author>
<author>
<name sortKey="Souiri, A" uniqKey="Souiri A">A Souiri</name>
</author>
<author>
<name sortKey="Kasmi, Y" uniqKey="Kasmi Y">Y Kasmi</name>
</author>
<author>
<name sortKey="Loutfi, C" uniqKey="Loutfi C">C Loutfi</name>
</author>
<author>
<name sortKey="Ennaji, Mm" uniqKey="Ennaji M">MM Ennaji</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Li, H" uniqKey="Li H">H Li</name>
</author>
<author>
<name sortKey="Yang, H" uniqKey="Yang H">H Yang</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Yan, S" uniqKey="Yan S">S Yan</name>
</author>
<author>
<name sortKey="Liu, X" uniqKey="Liu X">X Liu</name>
</author>
<author>
<name sortKey="Zhao, J" uniqKey="Zhao J">J Zhao</name>
</author>
<author>
<name sortKey="Xu, G" uniqKey="Xu G">G Xu</name>
</author>
<author>
<name sortKey="Zhao, Y" uniqKey="Zhao Y">Y Zhao</name>
</author>
<author>
<name sortKey="Zhang, G" uniqKey="Zhang G">G Zhang</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Khanh, Np" uniqKey="Khanh N">NP Khanh</name>
</author>
<author>
<name sortKey="Tan, Sw" uniqKey="Tan S">SW Tan</name>
</author>
<author>
<name sortKey="Yeap, Sk" uniqKey="Yeap S">SK Yeap</name>
</author>
<author>
<name sortKey="Lee, Hj" uniqKey="Lee H">HJ Lee</name>
</author>
<author>
<name sortKey="Choi, Ks" uniqKey="Choi K">KS Choi</name>
</author>
<author>
<name sortKey="Hair Bejo, M" uniqKey="Hair Bejo M">M Hair-Bejo</name>
</author>
<author>
<name sortKey="Bich, Tn" uniqKey="Bich T">TN Bich</name>
</author>
<author>
<name sortKey="Omar, Ar" uniqKey="Omar A">AR Omar</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Fulton, Rm" uniqKey="Fulton R">RM Fulton</name>
</author>
<author>
<name sortKey="Reed, Wm" uniqKey="Reed W">WM Reed</name>
</author>
<author>
<name sortKey="Thacker, Hl" uniqKey="Thacker H">HL Thacker</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Amarasinghe, A" uniqKey="Amarasinghe A">A Amarasinghe</name>
</author>
<author>
<name sortKey="Abdul Cader, Ms" uniqKey="Abdul Cader M">MS Abdul-Cader</name>
</author>
<author>
<name sortKey="Nazir, S" uniqKey="Nazir S">S Nazir</name>
</author>
<author>
<name sortKey="De Silva Senapathi, U" uniqKey="De Silva Senapathi U">U De Silva Senapathi</name>
</author>
<author>
<name sortKey="Van Der Meer, F" uniqKey="Van Der Meer F">F van der Meer</name>
</author>
<author>
<name sortKey="Cork, Sc" uniqKey="Cork S">SC Cork</name>
</author>
<author>
<name sortKey="Gomis, S" uniqKey="Gomis S">S Gomis</name>
</author>
<author>
<name sortKey="Abdul Careem, Mf" uniqKey="Abdul Careem M">MF Abdul-Careem</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Amarasinghe, A" uniqKey="Amarasinghe A">A Amarasinghe</name>
</author>
<author>
<name sortKey="Abdul Cader, Ms" uniqKey="Abdul Cader M">MS Abdul-Cader</name>
</author>
<author>
<name sortKey="Almatrouk, Z" uniqKey="Almatrouk Z">Z Almatrouk</name>
</author>
<author>
<name sortKey="Van Der Meer, F" uniqKey="Van Der Meer F">F van der Meer</name>
</author>
<author>
<name sortKey="Cork, Sc" uniqKey="Cork S">SC Cork</name>
</author>
<author>
<name sortKey="Gomis, S" uniqKey="Gomis S">S Gomis</name>
</author>
<author>
<name sortKey="Abdul Careem, Mf" uniqKey="Abdul Careem M">MF Abdul-Careem</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Da Silva, Sel" uniqKey="Da Silva S">SEL da Silva</name>
</author>
<author>
<name sortKey="Ferreira, Hl" uniqKey="Ferreira H">HL Ferreira</name>
</author>
<author>
<name sortKey="Garcia, Af" uniqKey="Garcia A">AF Garcia</name>
</author>
<author>
<name sortKey="Silva, Fes" uniqKey="Silva F">FES Silva</name>
</author>
<author>
<name sortKey="Gameiro, R" uniqKey="Gameiro R">R Gameiro</name>
</author>
<author>
<name sortKey="Fabri, Cuf" uniqKey="Fabri C">CUF Fabri</name>
</author>
<author>
<name sortKey="Vieira, Ds" uniqKey="Vieira D">DS Vieira</name>
</author>
<author>
<name sortKey="Cardoso, Tc" uniqKey="Cardoso T">TC Cardoso</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Han, X" uniqKey="Han X">X Han</name>
</author>
<author>
<name sortKey="Tian, Y" uniqKey="Tian Y">Y Tian</name>
</author>
<author>
<name sortKey="Guan, R" uniqKey="Guan R">R Guan</name>
</author>
<author>
<name sortKey="Gao, W" uniqKey="Gao W">W Gao</name>
</author>
<author>
<name sortKey="Yang, X" uniqKey="Yang X">X Yang</name>
</author>
<author>
<name sortKey="Zhou, L" uniqKey="Zhou L">L Zhou</name>
</author>
<author>
<name sortKey="Wang, H" uniqKey="Wang H">H Wang</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Kameka, Am" uniqKey="Kameka A">AM Kameka</name>
</author>
<author>
<name sortKey="Haddadi, S" uniqKey="Haddadi S">S Haddadi</name>
</author>
<author>
<name sortKey="Kim, Ds" uniqKey="Kim D">DS Kim</name>
</author>
<author>
<name sortKey="Cork, Sc" uniqKey="Cork S">SC Cork</name>
</author>
<author>
<name sortKey="Abdul Careem, Mf" uniqKey="Abdul Careem M">MF Abdul-Careem</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Meir, R" uniqKey="Meir R">R Meir</name>
</author>
<author>
<name sortKey="Maharat, O" uniqKey="Maharat O">O Maharat</name>
</author>
<author>
<name sortKey="Farnushi, Y" uniqKey="Farnushi Y">Y Farnushi</name>
</author>
<author>
<name sortKey="Simanov, L" uniqKey="Simanov L">L Simanov</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Breslin, Jj" uniqKey="Breslin J">JJ Breslin</name>
</author>
<author>
<name sortKey="Smith, Lg" uniqKey="Smith L">LG Smith</name>
</author>
<author>
<name sortKey="Fuller, Fj" uniqKey="Fuller F">FJ Fuller</name>
</author>
<author>
<name sortKey="Guy, Js" uniqKey="Guy J">JS Guy</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Klein, D" uniqKey="Klein D">D Klein</name>
</author>
</analytic>
</biblStruct>
<biblStruct>
<analytic>
<author>
<name sortKey="Reed, Lj" uniqKey="Reed L">LJ Reed</name>
</author>
<author>
<name sortKey="Muench, H" uniqKey="Muench H">H Muench</name>
</author>
</analytic>
</biblStruct>
</listBibl>
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<pmc-dir>properties open_access</pmc-dir>
<front>
<journal-meta>
<journal-id journal-id-type="nlm-ta">BMC Vet Res</journal-id>
<journal-id journal-id-type="iso-abbrev">BMC Vet. Res</journal-id>
<journal-title-group>
<journal-title>BMC Veterinary Research</journal-title>
</journal-title-group>
<issn pub-type="epub">1746-6148</issn>
<publisher>
<publisher-name>BioMed Central</publisher-name>
<publisher-loc>London</publisher-loc>
</publisher>
</journal-meta>
<article-meta>
<article-id pub-id-type="pmid">30526618</article-id>
<article-id pub-id-type="pmc">6288874</article-id>
<article-id pub-id-type="publisher-id">1720</article-id>
<article-id pub-id-type="doi">10.1186/s12917-018-1720-9</article-id>
<article-categories>
<subj-group subj-group-type="heading">
<subject>Research Article</subject>
</subj-group>
</article-categories>
<title-group>
<article-title>Comparative features of infections of two Massachusetts (Mass) infectious bronchitis virus (IBV) variants isolated from Western Canadian layer flocks</article-title>
</title-group>
<contrib-group>
<contrib contrib-type="author">
<name>
<surname>Amarasinghe</surname>
<given-names>Aruna</given-names>
</name>
<address>
<email>arunavetlk@gmail.com</email>
</address>
<xref ref-type="aff" rid="Aff1">1</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>De Silva Senapathi</surname>
<given-names>Upasama</given-names>
</name>
<address>
<email>yaseshwari.desilvase@ucalgary.ca</email>
</address>
<xref ref-type="aff" rid="Aff1">1</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Abdul-Cader</surname>
<given-names>Mohamed Sarjoon</given-names>
</name>
<address>
<email>mohamedsarjoon.moham@ucalgary.ca</email>
</address>
<xref ref-type="aff" rid="Aff1">1</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Popowich</surname>
<given-names>Shelly</given-names>
</name>
<address>
<email>shelly.popowich@usak.ca</email>
</address>
<xref ref-type="aff" rid="Aff2">2</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Marshall</surname>
<given-names>Frank</given-names>
</name>
<address>
<email>frankm@cable-lynx.net</email>
</address>
<xref ref-type="aff" rid="Aff3">3</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Cork</surname>
<given-names>Susan C.</given-names>
</name>
<address>
<email>sccork@ucalgary.ca</email>
</address>
<xref ref-type="aff" rid="Aff1">1</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>van der Meer</surname>
<given-names>Frank</given-names>
</name>
<address>
<email>fjvander@ucalgary.ca</email>
</address>
<xref ref-type="aff" rid="Aff1">1</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Gomis</surname>
<given-names>Susantha</given-names>
</name>
<address>
<email>smg127@mail.usask.ca</email>
</address>
<xref ref-type="aff" rid="Aff2">2</xref>
</contrib>
<contrib contrib-type="author" corresp="yes">
<contrib-id contrib-id-type="orcid">http://orcid.org/0000-0002-9974-7600</contrib-id>
<name>
<surname>Abdul-Careem</surname>
<given-names>Mohamed Faizal</given-names>
</name>
<address>
<phone>+403-220-4462</phone>
<email>faizal.abdulcareem@ucalgary.ca</email>
</address>
<xref ref-type="aff" rid="Aff1">1</xref>
</contrib>
<aff id="Aff1">
<label>1</label>
<institution-wrap>
<institution-id institution-id-type="ISNI">0000 0004 1936 7697</institution-id>
<institution-id institution-id-type="GRID">grid.22072.35</institution-id>
<institution>Department of Ecosystem and Public Health, Faculty of Veterinary Medicine,</institution>
<institution>University of Calgary, Health Research Innovation Center 2C53,</institution>
</institution-wrap>
3330 Hospital Drive NW, Calgary, AB T2N 4N1 Canada</aff>
<aff id="Aff2">
<label>2</label>
<institution-wrap>
<institution-id institution-id-type="ISNI">0000 0001 2154 235X</institution-id>
<institution-id institution-id-type="GRID">grid.25152.31</institution-id>
<institution>Department of Veterinary Pathology, Western College of Veterinary Medicine,</institution>
<institution>University of Saskatchewan,</institution>
</institution-wrap>
Saskatoon, SK S7N 5B5 Canada</aff>
<aff id="Aff3">
<label>3</label>
Marshall Swine and Poultry Health Services, 3831- Bay G- 44 Ave, Camrose, AB T4V 3T1 Canada</aff>
</contrib-group>
<pub-date pub-type="epub">
<day>10</day>
<month>12</month>
<year>2018</year>
</pub-date>
<pub-date pub-type="pmc-release">
<day>10</day>
<month>12</month>
<year>2018</year>
</pub-date>
<pub-date pub-type="collection">
<year>2018</year>
</pub-date>
<volume>14</volume>
<elocation-id>391</elocation-id>
<history>
<date date-type="received">
<day>9</day>
<month>7</month>
<year>2018</year>
</date>
<date date-type="accepted">
<day>27</day>
<month>11</month>
<year>2018</year>
</date>
</history>
<permissions>
<copyright-statement>© The Author(s). 2018</copyright-statement>
<license license-type="OpenAccess">
<license-p>
<bold>Open Access</bold>
This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (
<ext-link ext-link-type="uri" xlink:href="http://creativecommons.org/licenses/by/4.0/">http://creativecommons.org/licenses/by/4.0/</ext-link>
), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (
<ext-link ext-link-type="uri" xlink:href="http://creativecommons.org/publicdomain/zero/1.0/">http://creativecommons.org/publicdomain/zero/1.0/</ext-link>
) applies to the data made available in this article, unless otherwise stated.</license-p>
</license>
</permissions>
<abstract id="Abs1">
<sec>
<title>Background</title>
<p id="Par1">Infectious bronchitis virus (IBV) is one of the leading causes of mortality and morbidity in chickens. There are numerous serotypes and variants, which do not confer cross protection resulting in failure of currently used IBV vaccines. Although variant IBV isolates with major genetic differences have been subjected to comparative studies, it is unknown whether minor genetic differences in IBV variants within a serotype are different in terms of pathogenesis and eliciting host responses. Two Massachusetts (Mass) variant IBV isolates recovered from commercial layer flocks in the Western Canadian provinces of Alberta (AB) and Saskatchewan (SK) were compared genetically and evaluated for their pathogenicity, tissue distribution and ability to recruit and replicate in macrophages.</p>
</sec>
<sec>
<title>Results</title>
<p id="Par2">Although whole genome sequencing of these two Mass IBV isolates showed low similarity with the M41 vaccinal strain, they had an identical nucleotide sequence at open reading frames (ORFs) 3a, 3b, envelop (E), matrix (M), 5a and 5b. The rest of the ORFs of these 2 IBV isolates showed 99.9% nucleotide similarity. However, upon experimental infection, we found that the IBV isolate originating from AB was different to the one that originated in SK due to higher tracheal lesion scores and lower lung viral replication and lower genome loads in cecal tonsils. Nevertheless, both IBV isolates elicited host responses characterized by significant macrophage recruitment to the respiratory tract and there was evidence that both IBV isolates replicated within tracheal and lung macrophages.</p>
</sec>
<sec>
<title>Conclusions</title>
<p id="Par3">Overall, this study shows that Mass variant IBV isolates, although possessing minor genetic variations, can lead to significant differences in pathogenicity in young chickens. Further studies are required to investigate the pathogenicity of these two Mass variant IBV isolates in laying hens.</p>
</sec>
<sec>
<title>Electronic supplementary material</title>
<p>The online version of this article (10.1186/s12917-018-1720-9) contains supplementary material, which is available to authorized users.</p>
</sec>
</abstract>
<kwd-group xml:lang="en">
<title>Keywords</title>
<kwd>Infectious bronchitis virus</kwd>
<kwd>Whole genome sequencing</kwd>
<kwd>Tissue distribution</kwd>
<kwd>Pathogenicity</kwd>
<kwd>Macrophage response</kwd>
</kwd-group>
<funding-group>
<award-group>
<funding-source>
<institution-wrap>
<institution-id institution-id-type="FundRef">http://dx.doi.org/10.13039/100008921</institution-id>
<institution>Ministry of Agriculture - Saskatchewan</institution>
</institution-wrap>
</funding-source>
<award-id>20140185</award-id>
<principal-award-recipient>
<name>
<surname>Gomis</surname>
<given-names>Susantha</given-names>
</name>
</principal-award-recipient>
</award-group>
</funding-group>
<funding-group>
<award-group>
<funding-source>
<institution>Saskatchewan Eggs Producers</institution>
</funding-source>
<award-id>20140185</award-id>
<principal-award-recipient>
<name>
<surname>Gomis</surname>
<given-names>Susantha</given-names>
</name>
</principal-award-recipient>
</award-group>
</funding-group>
<custom-meta-group>
<custom-meta>
<meta-name>issue-copyright-statement</meta-name>
<meta-value>© The Author(s) 2018</meta-value>
</custom-meta>
</custom-meta-group>
</article-meta>
</front>
<body>
<sec id="Sec1">
<title>Background</title>
<p id="Par4">Infectious bronchitis virus (IBV) belongs to the family
<italic>Coronaviridae</italic>
. Traditionally, IBV is considered to be a host-specific respiratory pathogen in chickens and IBV initially replicates at the route of entry, the tracheal mucosa [
<xref ref-type="bibr" rid="CR1">1</xref>
,
<xref ref-type="bibr" rid="CR2">2</xref>
]. However, identification of new variants and/or serotypes of IBV have shown a wide variation of tissue tropism including urinary [
<xref ref-type="bibr" rid="CR3">3</xref>
<xref ref-type="bibr" rid="CR10">10</xref>
], gastrointestinal [
<xref ref-type="bibr" rid="CR6">6</xref>
,
<xref ref-type="bibr" rid="CR9">9</xref>
,
<xref ref-type="bibr" rid="CR11">11</xref>
,
<xref ref-type="bibr" rid="CR12">12</xref>
], oviduct [
<xref ref-type="bibr" rid="CR9">9</xref>
,
<xref ref-type="bibr" rid="CR13">13</xref>
] and bursa of Fabricius [
<xref ref-type="bibr" rid="CR11">11</xref>
,
<xref ref-type="bibr" rid="CR14">14</xref>
]. IBV is known to replicate in the reproductive tract epithelium in layers leading to reduced egg production and shell defective eggs [
<xref ref-type="bibr" rid="CR15">15</xref>
,
<xref ref-type="bibr" rid="CR16">16</xref>
]. False layer syndrome, which is associated with cystic oviduct formation occurs with IBV infection in early life [
<xref ref-type="bibr" rid="CR17">17</xref>
,
<xref ref-type="bibr" rid="CR18">18</xref>
]. IBV can also replicate in the testes of cockerels [
<xref ref-type="bibr" rid="CR19">19</xref>
].</p>
<p id="Par5">An array of serotypes and strains of IBV infect chickens throughout the world [
<xref ref-type="bibr" rid="CR2">2</xref>
]. Genetic events such as insertion(s) and deletions [
<xref ref-type="bibr" rid="CR20">20</xref>
,
<xref ref-type="bibr" rid="CR21">21</xref>
], point mutations [
<xref ref-type="bibr" rid="CR22">22</xref>
], and recombination [
<xref ref-type="bibr" rid="CR23">23</xref>
<xref ref-type="bibr" rid="CR27">27</xref>
] contribute to genomic variations of IBV [
<xref ref-type="bibr" rid="CR28">28</xref>
]. The spike 1 (S1) gene is highly variable among IBV strains and it encodes epitopes, which bind to neutralizing antibodies [
<xref ref-type="bibr" rid="CR29">29</xref>
]. A change in the amino acid sequence as small as 2 to 3% in the S1 subunit can result in changes in the antigenicity of the virus [
<xref ref-type="bibr" rid="CR30">30</xref>
]. Based on these changes of the S1 protein, numerous IBV strains have been characterized throughout the world [
<xref ref-type="bibr" rid="CR1">1</xref>
]. Therefore, either the partial [
<xref ref-type="bibr" rid="CR31">31</xref>
<xref ref-type="bibr" rid="CR33">33</xref>
] or the full-length [
<xref ref-type="bibr" rid="CR34">34</xref>
<xref ref-type="bibr" rid="CR39">39</xref>
] of the S1 glycoprotein gene has been used in the molecular characterization of IBV isolates. However, using whole genome sequencing it has been observed that genes other than S1 may play a role in the pathogenicity of IBV infection [
<xref ref-type="bibr" rid="CR40">40</xref>
,
<xref ref-type="bibr" rid="CR41">41</xref>
]. Currently, there are no IBV reference full genome sequences available for Canadian IBV isolates but partial [
<xref ref-type="bibr" rid="CR33">33</xref>
] or complete [
<xref ref-type="bibr" rid="CR37">37</xref>
] S1 sequences are available.</p>
<p id="Par6">Previously, comparative studies have been conducted to elucidate differential pathogenicity and host responses between variant IBV isolates with larger genome variability such as nephropathogenic and Massachusetts (Mass) IBV isolates recovered from various geographical areas [
<xref ref-type="bibr" rid="CR18">18</xref>
,
<xref ref-type="bibr" rid="CR42">42</xref>
<xref ref-type="bibr" rid="CR46">46</xref>
]. Since it is well known that very minor changes in the genome of viruses [
<xref ref-type="bibr" rid="CR47">47</xref>
] including IBV [
<xref ref-type="bibr" rid="CR48">48</xref>
,
<xref ref-type="bibr" rid="CR49">49</xref>
] can lead to difference in pathogenicity, comparative studies involving variant IBV isolates are required.</p>
<p id="Par7">A wide array of IBV variants are affecting commercial broiler, layer and breeder flocks in Canada [
<xref ref-type="bibr" rid="CR33">33</xref>
,
<xref ref-type="bibr" rid="CR37">37</xref>
,
<xref ref-type="bibr" rid="CR50">50</xref>
,
<xref ref-type="bibr" rid="CR51">51</xref>
]. For example, Mass type IBV variants are impacting the commercial egg production in Western Canada [
<xref ref-type="bibr" rid="CR50">50</xref>
]. This study showed that Mass type IBV infection of 27-week old layers at peak production, lead to loss of egg production for about 2 weeks followed by production of shell less, small and defective eggs for another 2 days before the egg production bounced back to normal production. The duration of the loss of marketable eggs appears to be 16 days. In an 8000 bird layer flock of Western Canada, we observed 47.6% drop in egg production for 10 days and we isolated a Mass type variant IBV from this flock. Based on Egg Farmers of Alberta’s price of $2.15/dozen of large eggs, the loss for 10 days is calculated to be $6823 for this particular outbreak and the loss will be higher if the infection persists beyond 10 days or reoccur. In addition to this direct loss of egg production, egg grading stations need to outsource supply of eggs during these production drops and also some of the affected birds may succumb to secondary bacterial infections without a recovery, hence the indirect losses are substantially higher than the direct costs of IBV infection.</p>
<p id="Par8">The present study was conducted with the aim to elucidate the genetic differences of two Mass variant IBV isolates (15AB-01 and 15SK-02) that originated from layer flocks in Western Canada, Alberta (AB) and Saskatchewan (SK). Additionally, we aimed to describe whether these genetic changes in Mass isolates 15AB-01 and 15SK-02 lead to differential pathogenicity, tissue distribution and macrophage responses in the respiratory tract.</p>
</sec>
<sec id="Sec2">
<title>Results</title>
<sec id="Sec3">
<title>Molecular characterization of the mass IBV isolates, 15AB-01 and 15SK-02</title>
<p id="Par9">For the isolate 15AB-01, total of 525,998 and 908,860 read sets were obtained from the direct allantoic fluid sample and layered virus sample, respectively from the Illumina MiSeq workflow. From these, a total of 418,446 and 825,147 reads were aligned with the reference IBV genomes. For isolate 15SK-02, a total of 4,355,902 and 3,731,578 reads were obtained from the direct allantoic fluid sample and layered virus sample, respectively, of which, a total of 3,146,584 and 3,152,622 reads were aligned with the IBV reference genomes. The number of reads obtained from Illumina MiSeq for 15SK-02 was 10 times higher than for 15AB-01 and the viral genome concentration of the submitted complimentary (c)DNA was also much higher for 15SK-02 than for 15AB-01 (real-time PCR cycle threshold or Ct value difference of 3.54). Entirely based on the NGS output, the complete genomes obtained for the Mass IBV isolates, 15AB-01 and 15SK-02 were 27.6 kb in length. As determined by the open reading frames (ORF) predictor, both isolates had similar genome organization with 11 ORFs and they were 5′-1a-1b-S-3a-3b-E-M-5a-5b-N-3′ (Table 
<xref rid="Tab1" ref-type="table">1</xref>
). Between the isolates, 15AB-01 and 15SK-02, there was 99.9% nucleotide similarity. With the M41 vaccine strain, 15AB-01 and 15SK-02 shared a genetic similarity of 92.8 and 92.7%, respectively. 15AB-01 and 15SK-02 Mass IBV isolates shared 92.8 and 92.7% nucleotide similarity with the M41 vaccine strain (GQ504725), respectively. Nucleotide similarities between Mass IBV isolates, 15AB-01 and 15SK-02 across the ORFs 3a, 3b, E, M, 5a and 5b were 100%. The remainder of genes were over 99% identical (Table 
<xref rid="Tab2" ref-type="table">2</xref>
). Nucleotide similarities between Mass IBV isolates, 15AB-01 and 15SK-02 across the hypervariable S gene regions, S1 and S2, respectively were 99.6 and 99.8%. The lowest nucleotide similarity these two isolates shared with the vaccine M41 strain was 86.4% in the ORF 5a and the highest similarity was 100% in the ORF M. The S1 and S2 identities of these two IBV isolates with the M41 vaccine strain were 95.5 and 97.6%, respectively (Table
<xref rid="Tab2" ref-type="table">2</xref>
). Additionally, our IBV isolates, 15AB-01 and 15SK-02 were compared with important reference strains in Mass type cluster (Table 
<xref rid="Tab3" ref-type="table">3</xref>
) and they are most variable from the Beaudette IBV strain and very closely related to other Mass type IBV strains.
<table-wrap id="Tab1">
<label>Table 1</label>
<caption>
<p>Genes, coding regions, and deduced proteins of the current Mass IBV isolates, 15AB-01 and 15SK-02</p>
</caption>
<table frame="hsides" rules="groups">
<thead>
<tr>
<th>Gene</th>
<th>Translation Frame</th>
<th>Gene location</th>
<th>Nucleotide length (bp)</th>
<th>Number of amino acids</th>
</tr>
</thead>
<tbody>
<tr>
<td>1a</td>
<td>1</td>
<td>529–12,390</td>
<td>11,862</td>
<td>3953</td>
</tr>
<tr>
<td>1b</td>
<td>3</td>
<td>12,465–20,423</td>
<td>7959</td>
<td>2652</td>
</tr>
<tr>
<td>S</td>
<td>1</td>
<td>20,374–23,862</td>
<td>3489</td>
<td>1162</td>
</tr>
<tr>
<td>3a</td>
<td>3</td>
<td>23,862–24,035</td>
<td>174</td>
<td>57</td>
</tr>
<tr>
<td>3b</td>
<td>2</td>
<td>24,035–24,229</td>
<td>195</td>
<td>64</td>
</tr>
<tr>
<td>E</td>
<td>3</td>
<td>24,210–24,539</td>
<td>330</td>
<td>109</td>
</tr>
<tr>
<td>M</td>
<td>1</td>
<td>24,511–25,188</td>
<td>678</td>
<td>225</td>
</tr>
<tr>
<td>5a</td>
<td>3</td>
<td>25,539–25,736</td>
<td>198</td>
<td>65</td>
</tr>
<tr>
<td>5b</td>
<td>2</td>
<td>25,733–25,981</td>
<td>249</td>
<td>82</td>
</tr>
<tr>
<td>N</td>
<td>1</td>
<td>25,924–27,153</td>
<td>1230</td>
<td>409</td>
</tr>
</tbody>
</table>
</table-wrap>
<table-wrap id="Tab2">
<label>Table 2</label>
<caption>
<p>Nucleotide similarity of 15AB-01 and 15SK-02 across various genes with M41 vaccine strain GQ504725</p>
</caption>
<table frame="hsides" rules="groups">
<thead>
<tr>
<th>Strain</th>
<th>ORF1a</th>
<th>ORF 1b</th>
<th>S1</th>
<th>S2</th>
<th>ORF3a</th>
<th>ORF3b</th>
<th>E</th>
<th>M</th>
<th>ORF5a</th>
<th>ORF5b</th>
<th>N</th>
</tr>
</thead>
<tbody>
<tr>
<td>15AB-01</td>
<td>90.7</td>
<td>96.8</td>
<td>95.5</td>
<td>97.6</td>
<td>91.2</td>
<td>96.9</td>
<td>94.5</td>
<td>100</td>
<td>86.4</td>
<td>89.2</td>
<td>94.1</td>
</tr>
<tr>
<td>15SK-02</td>
<td>90.6</td>
<td>96.8</td>
<td>95.5</td>
<td>97.9</td>
<td>91.2</td>
<td>96.9</td>
<td>94.5</td>
<td>100</td>
<td>86.4</td>
<td>89.2</td>
<td>94.5</td>
</tr>
<tr>
<td>
<sup>a</sup>
15AB-01 Vs 15SK-02</td>
<td>99.9</td>
<td>99.9</td>
<td>99.6</td>
<td>99.8</td>
<td>100</td>
<td>100</td>
<td>100</td>
<td>100</td>
<td>100</td>
<td>100</td>
<td>99.5</td>
</tr>
</tbody>
</table>
<table-wrap-foot>
<p>
<sup>a</sup>
Third raw depicts the nucleotide similarities between current Mass IBV isolates 15AB-01 and 15SK-02 across various genes</p>
</table-wrap-foot>
</table-wrap>
<table-wrap id="Tab3">
<label>Table 3</label>
<caption>
<p>Nucleotide identities of 15AB-01 and 15SK-02 across various genes with Mass-type reference strains</p>
</caption>
<table frame="hsides" rules="groups">
<thead>
<tr>
<th>Strain</th>
<th>ORF 1a</th>
<th>ORF 1b</th>
<th>S1</th>
<th>S2</th>
<th>ORF 3a</th>
<th>ORF 3b</th>
<th>E</th>
<th>M</th>
<th>ORF 5a</th>
<th>ORF 5b</th>
<th>N</th>
</tr>
</thead>
<tbody>
<tr>
<td>Beaudette</td>
<td>91.5/91.5</td>
<td>99.9/93.9</td>
<td>97.8/97.8</td>
<td>98.1/98.2</td>
<td>97.7/97.1</td>
<td>98.2/98.2</td>
<td>98.7/98.7</td>
<td>98.8/98.8</td>
<td>98.9/98.9</td>
<td>90.8/90.8</td>
<td>92.6/92.7</td>
</tr>
<tr>
<td>TN92–03 (India)</td>
<td>93.2/93.2</td>
<td>94.0/93.9</td>
<td>97.2/97.0</td>
<td>98.3/98.4</td>
<td>97.7/98.3</td>
<td>97.0/97.0</td>
<td>98.4/98.4</td>
<td>99.0/99.0</td>
<td>100/100</td>
<td>95.6/95.6</td>
<td>97.6/99.8</td>
</tr>
<tr>
<td>Mass41/1972 (USA)</td>
<td>99.7/99.7</td>
<td>99.5/99.5</td>
<td>99.8/99.8</td>
<td>99.2/99.3</td>
<td>98.8/99.4</td>
<td>100/100</td>
<td>99.7/99.7</td>
<td>99.0/99.0</td>
<td>100/100</td>
<td>100/100</td>
<td>99.6/99.8</td>
</tr>
<tr>
<td>Mass/USA (AY851295)</td>
<td>100/99.9</td>
<td>99.9/99.9</td>
<td>99.9/99.9</td>
<td>99.9/99.9</td>
<td>100/99.4</td>
<td>100/100</td>
<td>99.7/99.7</td>
<td>100/100</td>
<td>100/100</td>
<td>100/100</td>
<td>99.8/99.6</td>
</tr>
<tr>
<td>ck/CH/LHLJ/091205 (China)</td>
<td>99.9/99.9</td>
<td>99.9/99.9</td>
<td>99.9/99.9</td>
<td>99.9/99.9</td>
<td>100/99.4</td>
<td>100/100</td>
<td>100/100</td>
<td>100/100</td>
<td>100/100</td>
<td>100/100</td>
<td>100/99.8</td>
</tr>
<tr>
<td>M41/USA (DQ834384)</td>
<td>100/100</td>
<td>100/99.9</td>
<td>99.9/99.9</td>
<td>99.7/99.7</td>
<td>100/99.4</td>
<td>100/100</td>
<td>100/100</td>
<td>99.9/99.9</td>
<td>100/100</td>
<td>100/100</td>
<td>99.7/99.8</td>
</tr>
<tr>
<td>Mass/USA (GQ504724)</td>
<td>100/100</td>
<td>99.9/99.9</td>
<td>99.9/99.9</td>
<td>99.8/99.8</td>
<td>100/99.4</td>
<td>100/100</td>
<td>100/100</td>
<td>100/100</td>
<td>100/100</td>
<td>99.6/99.6</td>
<td>99.8/99.8</td>
</tr>
<tr>
<td>M41/1965 (USA)</td>
<td>99.9/99.9</td>
<td>100/100</td>
<td>99.9/99.9</td>
<td>99.8/99.8</td>
<td>98.8/99.4</td>
<td>100/100</td>
<td>100/100</td>
<td>100/100</td>
<td>100/100</td>
<td>100/100</td>
<td>99.6/99.8</td>
</tr>
<tr>
<td>M41/1985 (USA)</td>
<td>100/100</td>
<td>99.9/100</td>
<td>99.9/100</td>
<td>99.8/99.8</td>
<td>98.8/99.4</td>
<td>100/100</td>
<td>100/100</td>
<td>100/100</td>
<td>100/100</td>
<td>100/100</td>
<td>99.7/99.8</td>
</tr>
</tbody>
</table>
<table-wrap-foot>
<p>In each cell of the table, the first value is the nucleotide identity shared with the IBV isolate 15AB-01 and the second value is the nucleotide identity shared with the IBV isolate 15SK-02</p>
</table-wrap-foot>
</table-wrap>
</p>
<p id="Par10">Full genome nucleotide distance matrix and heat maps of these isolates with 23 selected reference sequences are shown in the Fig. 
<xref rid="Fig1" ref-type="fig">1</xref>
. The highest similarity of 15AB-01 and 15SK-02 shared with M41 strain (USA) was 99.9%. The lowest similarity was 87.5% with the United Kingdom (UK) strain 1148 (Fig.
<xref rid="Fig1" ref-type="fig">1</xref>
). Phylogenetic reconstruction with reference IBV strains based on the full genome sequence revealed a close relation with two IBV isolates to the USA Mass type virus (Fig. 
<xref rid="Fig2" ref-type="fig">2</xref>
). The isolate 15AB-01 appeared to be more recently evolved than the 15SK-02 isolate. However, clear separation of these two IBV isolates (cluster M1) from the vaccine M41 strain (Cluster M11) is evident (Fig.
<xref rid="Fig2" ref-type="fig">2</xref>
).
<fig id="Fig1">
<label>Fig. 1</label>
<caption>
<p>Nucleotide similarity matrix and heat maps of 23-selected reference strains and 2 current Mass IBV isolates. Full genome sequences of Western Canadian isolates (15AB-01 and 15SK-02) were aligned with 23 reference sequences retrieved from the GenBank. The resulting genetic distance matrix along with heat maps based on the similarity index is shown</p>
</caption>
<graphic xlink:href="12917_2018_1720_Fig1_HTML" id="MO1"></graphic>
</fig>
<fig id="Fig2">
<label>Fig. 2</label>
<caption>
<p>Phylogenetic tree of 45 IBV whole genome sequences including Western Canadian Mass IBV isolates. A maximum likely hood phylogenetic tree was inferred based on the whole genomes of 15AB-01 and 15SK-02 isolates of IBV and 43 IBV genome sequences retrieved from GenBank using RAxML with GTR GAMMA nucleotide substitution model with 1000 bootstrap iterations</p>
</caption>
<graphic xlink:href="12917_2018_1720_Fig2_HTML" id="MO2"></graphic>
</fig>
</p>
</sec>
<sec id="Sec4">
<title>Clinical observations and body weight changes following IBV infection of chickens</title>
<p id="Par11">From 2 to 8 days post-infection (dpi), there were two birds with mildly increased respiration (more labored and higher frequency) from the IBV (15AB-01) infected group. On 9 and 10 dpi, there were 3 birds with mildly increased breathing. Only one bird continued to show a mild breathing increase by 11 dpi and thereafter all birds were bright and alert. In contrast, 2 birds from the 15SK-02 infected group started showing increased breathing as late as 7 dpi and continued until 11 dpi. The control birds were alert and active throughout. The highest body weight gain was achieved by the uninfected control group followed by the group infected with the isolate 15AB-01 and 15SK-02 (Fig. 
<xref rid="Fig3" ref-type="fig">3</xref>
). However, the difference in bodyweights was not significant (
<italic>p</italic>
 > 0.05).
<fig id="Fig3">
<label>Fig. 3</label>
<caption>
<p>Bodyweight changes following infection with Western Canadian Mass-type IBV isolates. The average body weight of each group (IBV 15AB-01, IBV 15SK-02 and uninfected control) of chickens at different time points are depicted in the graph. The error bars represent the ±standard error of mean (SEM)</p>
</caption>
<graphic xlink:href="12917_2018_1720_Fig3_HTML" id="MO3"></graphic>
</fig>
</p>
</sec>
<sec id="Sec5">
<title>Histopathological changes following IBV infection of chickens</title>
<p id="Par12">Histopathological examination of both trachea and lung at 4 dpi showed lesions suggestive of infectious bronchitis. In the trachea, these histological changes included de-ciliation of tracheal mucosal cells, lymphoid infiltration into mucosa and underlying submucosa, and mucosal detachment from the lamina propria (Fig. 
<xref rid="Fig4" ref-type="fig">4</xref>
a). Squamous metaplasia of the tracheal mucosa was prominent in trachea of both IBV infected groups (Fig.
<xref rid="Fig4" ref-type="fig">4</xref>
a). Average lesion score of the trachea infected with the isolate 15AB-01 (
<italic>p</italic>
 < 0.001) and 15SK-02 (
<italic>p</italic>
 < 0.01) were significantly higher than that of trachea from uninfected controls (Fig.
<xref rid="Fig4" ref-type="fig">4</xref>
b). Also lesion scores of the trachea from 15AB-01 infected chicken were significantly higher than that of trachea from chicken infected with isolate 15SK-02 (
<italic>p</italic>
 < 0.01) (Fig.
<xref rid="Fig4" ref-type="fig">4</xref>
b). In the lungs, mononuclear cell infiltration and hemorrhage into the parabronchial lumen were observed in the infected lungs compared to the uninfected controls (Fig.
<xref rid="Fig4" ref-type="fig">4</xref>
a). We observed variation in lung lesion scores within groups, yet significant outliers were not detected when the data were subjected to Grubbs test. The lung lesion scores of the lung from 15AB-01 infected chicken were significantly higher than that of the lungs from uninfected controls. However, the lung lesion score between the lung from 15SK-02 infected chicken and the uninfected controls was not statistically significant (
<italic>p</italic>
 > 0.05) (Fig.
<xref rid="Fig4" ref-type="fig">4</xref>
c).
<fig id="Fig4">
<label>Fig. 4</label>
<caption>
<p>Histological changes in trachea and lungs induced by Western Canadian Mass-type IBV isolates. The representative images of trachea and lungs (
<bold>a</bold>
) and average lesion scores of trachea (
<bold>b</bold>
) and lung (
<bold>c</bold>
) for each group of chickens (IBV 15AB-01, IBV 15SK-02 and uninfected control) at 4 dpi are depicted. a = cilia, b = tracheal cartilage, c = tracheal lumen, d = tracheal mucosa, e = parabronchial lumen and f = inter-parabronchial septum. The error bars represent the ±SEM. Statistical significance: *
<italic>p</italic>
 < 0.05, **
<italic>p</italic>
 < 0.01, ***
<italic>p</italic>
 < 0.001</p>
</caption>
<graphic xlink:href="12917_2018_1720_Fig4_HTML" id="MO4"></graphic>
</fig>
</p>
</sec>
<sec id="Sec6">
<title>IBV genome loads in oropharyngeal and cloacal swabs and tissue samples</title>
<p id="Par13">The average genome loads on oropharyngeal swabs were higher than the genome loads on cloacal swabs for both Mass IBV isolates at all time points (Fig. 
<xref rid="Fig5" ref-type="fig">5</xref>
a and b). In the trachea, a genome load of 1 × 10
<sup>6</sup>
up to 3 dpi could be detected in chicken infected with either isolate. By 12 dpi, the genome loads in the trachea declined by 2-folds. The genome load in the cloaca was consistent for the 15AB-01 throughout the observation period. However, in the cloaca of birds infected with 15SK-02 no IBV genome at 12 dpi could be detected (Fig.
<xref rid="Fig5" ref-type="fig">5</xref>
b). No IBV genome load was detected in the uninfected controls (results not shown).
<fig id="Fig5">
<label>Fig. 5</label>
<caption>
<p>IBV genome loads in swabs and tissues following infection with Western Canadian Mass-type IBV isolates. For each time point, the average log
<sub>10</sub>
starting IBV copies per 200 ng of extracted RNA from the oropharyngeal (
<bold>a</bold>
) and cloacal (
<bold>b</bold>
) swabs and various tissues (
<bold>c</bold>
) at 4 and 14 dpi are shown with the SEM. Statistical significance: *
<italic>p</italic>
 < 0.05, **
<italic>p</italic>
 < 0.01, ***
<italic>p</italic>
 < 0.001</p>
</caption>
<graphic xlink:href="12917_2018_1720_Fig5_HTML" id="MO5"></graphic>
</fig>
</p>
<p id="Par14">The highest genome loads in tissue were found in trachea and lung tissues of chicken infected with either isolate at 4 dpi, however, the genome load in lung tissue was significantly lower for IBV isolate 15AB-01 (Fig.
<xref rid="Fig5" ref-type="fig">5</xref>
c). At 4 and 14 dpi, the genome loads in trachea were similar for IBV isolates (15AB-01 and 15SK-02). By 14 dpi, tracheal and lung genome loads fell considerably for both isolates. Cecal tonsil genome load was declined by 14 dpi only in birds infected with the isolate 15AB-01 and the difference was statistically significant (
<italic>p</italic>
 < 0.01). There was no difference in genome loads in the kidney tissue between two time points in both strains. The uterine genome loads were available only at 14 dpi for both strains. None of the uninfected control birds had detectable IBV genome loads (results not shown).</p>
</sec>
<sec id="Sec7">
<title>Immunofluorescence data for IBV antigens and macrophages at 4 dpi</title>
<p id="Par15">Similar to IBV genome loads, we did not see a difference in IBV antigen expression in trachea of 15AB-01 and 15SK-02 infected chickens (
<italic>p</italic>
 > 0.05, Fig. 
<xref rid="Fig6" ref-type="fig">6</xref>
b). Similar to the IBV genome load, the amount of IBV antigen was significantly lower in 15AB-01 infected lung than that of 15SK-02 infected lung (
<italic>p</italic>
 < 0.01, Fig. 
<xref rid="Fig7" ref-type="fig">7</xref>
b). The lung and tracheal macrophage numbers were not significantly different in the two groups of experimentally infected chicken, indicating a similar response to the two strains of Mass IBV used in the study (
<italic>p</italic>
 > 0.05). Macrophage infiltration was significantly higher (
<italic>p</italic>
 < 0.05) in tracheas infected with both Mass IBV isolates compared to the tracheas of uninfected controls (Fig.
<xref rid="Fig6" ref-type="fig">6</xref>
a and c). Similarly, lungs infected with either Mass IBV isolates had significantly higher number of macrophages (
<italic>p</italic>
 < 0.01) than lungs from uninfected controls (Fig.
<xref rid="Fig7" ref-type="fig">7</xref>
a and c). In both the trachea and lung, there was no significant difference in IBV positive macrophages between two Mass IBV isolates (Figs. 
<xref rid="Fig6" ref-type="fig">6</xref>
d and
<xref rid="Fig7" ref-type="fig">7</xref>
d).
<fig id="Fig6">
<label>Fig. 6</label>
<caption>
<p>Mass-type IBV isolates recovered from Western Canadian layer flocks increase macrophage numbers in trachea. The representative images of trachea stained for IBV antigens and macrophages (
<bold>a</bold>
) and quantitative data for IBV antigens (
<bold>b</bold>
), macrophages (
<bold>c</bold>
) and IBV antigen positive macrophages (
<bold>d</bold>
) at 4 dpi are depicted. The error bars represent the ± SEM. Statistical significance: *
<italic>p</italic>
 < 0.05, **
<italic>p</italic>
 < 0.01, ***
<italic>p</italic>
 < 0.001</p>
</caption>
<graphic xlink:href="12917_2018_1720_Fig6_HTML" id="MO6"></graphic>
</fig>
<fig id="Fig7">
<label>Fig. 7</label>
<caption>
<p>Mass-type IBV isolates recovered from Western Canadian layer flocks increase macrophage numbers in lungs. The representative images of lungs stained for IBV antigens and macrophages (
<bold>a</bold>
) and quantitative data for IBV antigens (
<bold>b</bold>
), macrophages (
<bold>c</bold>
) and IBV antigen positive macrophages (
<bold>d</bold>
) at 4 dpi are depicted. The error bars represent the ± SEM. Statistical significance: *
<italic>p</italic>
 < 0.05, **
<italic>p</italic>
 < 0.01, ***
<italic>p</italic>
 < 0.001</p>
</caption>
<graphic xlink:href="12917_2018_1720_Fig7_HTML" id="MO7"></graphic>
</fig>
</p>
</sec>
</sec>
<sec id="Sec8">
<title>Discussion</title>
<p id="Par16">The work described in this manuscript focused, first, to see the genetic differences of two Mass variant IBV isolates originating from layer flocks in two provinces of Western Canada. Second, we evaluated if infection of SPF chicken with genetically different Mass IBV variants would result in a different pathogenicity, tissue distribution and macrophage responses in the respiratory tract. Since the S1 protein is the main inducer of virus-neutralizing antibodies [
<xref ref-type="bibr" rid="CR52">52</xref>
<xref ref-type="bibr" rid="CR54">54</xref>
], either partial [
<xref ref-type="bibr" rid="CR31">31</xref>
<xref ref-type="bibr" rid="CR33">33</xref>
] or full-length [
<xref ref-type="bibr" rid="CR34">34</xref>
<xref ref-type="bibr" rid="CR39">39</xref>
] S1 glycoprotein gene have been used in many studies involving IBV molecular characterization throughout the world including previous Canadian studies. Using whole genome sequencing it has been shown that genes other than S1 may be playing roles in the pathogenicity of IBV [
<xref ref-type="bibr" rid="CR41">41</xref>
,
<xref ref-type="bibr" rid="CR55">55</xref>
]. To our knowledge this is the first study, which investigated the full genome of Canadian IBV isolates. Although, IBV strains circulating in other countries have been characterized and changes in the IBV genome due to mutations and recombination have been shown [
<xref ref-type="bibr" rid="CR40">40</xref>
,
<xref ref-type="bibr" rid="CR55">55</xref>
<xref ref-type="bibr" rid="CR59">59</xref>
], our whole genome sequencing data shows that mutations are more common than the recombination and genes other than S1 have been affected by mutations. These changes in the genome of these two IBV isolates reflected in the differences in the pathogenicity although host response characterized by macrophage recruitment into the respiratory tract were similar.</p>
<p id="Par17">Previously, studies have been conducted to compare pathogenicity and immune responses between nephropathogenic (i.e. QX like IBV strains) and Mass type IBV isolates recovered from various geographical areas [
<xref ref-type="bibr" rid="CR18">18</xref>
,
<xref ref-type="bibr" rid="CR42">42</xref>
,
<xref ref-type="bibr" rid="CR43">43</xref>
,
<xref ref-type="bibr" rid="CR45">45</xref>
,
<xref ref-type="bibr" rid="CR46">46</xref>
]. Nephropathogenic QX-type strains are distantly related to the M41 strain and their genetic similarity is 78.6% [
<xref ref-type="bibr" rid="CR60">60</xref>
]. Based on the partial S1 sequence analysis of three nephropathogenic IBV isolates, which belong to the Mass genotype revealed that they are more than 99.1 and 97.8% identical to the classical M41 strain in nucleotide and deduced amino acid sequences, respectively [
<xref ref-type="bibr" rid="CR61">61</xref>
]. Since it is well known that very minor changes in the genome of viruses [
<xref ref-type="bibr" rid="CR47">47</xref>
] including IBV [
<xref ref-type="bibr" rid="CR49">49</xref>
,
<xref ref-type="bibr" rid="CR62">62</xref>
] lead to differences in pathogenicity and host responses, we chose two Mass type IBV isolates expected to have minor genome differences for evaluation of pathogenicity, tissue distribution and host responses. Mass type IBV isolates have been shown to replicate in the respiratory tissues, kidney, cecal tonsils and gastrointestinal tissues [
<xref ref-type="bibr" rid="CR18">18</xref>
,
<xref ref-type="bibr" rid="CR42">42</xref>
,
<xref ref-type="bibr" rid="CR43">43</xref>
]. In agreement with these studies, we also observed that Mass IBV isolates originating from Western Canadian layer flocks replicate in tissues other than respiratory tissues. In our study, Mass IBV isolate originating in SK had significantly higher genome load in the lungs than the one originating in AB, though there was no difference of lung lesion score between these two IBV isolates. On the other hand, in trachea, the genome loads of these two Mass IBV isolates were not different, although the Mass IBV isolate originated in AB had significantly higher lesion scores. It is also noteworthy that Mass IBV isolate originating in SK was not detectable at 12 dpi in cloacal swabs. Since the major source of IBV genome in cloaca is due to IBV replication in gastrointestinal tract [
<xref ref-type="bibr" rid="CR63">63</xref>
], it is possible that 15SK-02 IBV infection in gastrointestinal tract may have been cleared by 12 dpi. Overall, these two Mass IBV variants have similar pathogenicity with only slight differences which also agree with the minor genetic differences of these 2 Mass type IBV isolates.</p>
<p id="Par18">Literature that describe pathogenicity using Canadian IBV isolates are scarce [
<xref ref-type="bibr" rid="CR8">8</xref>
] Although this study evaluated the pathogenicity of 5 IBV isolates originated in Ontario poultry flocks, IBV shedding, tissue distribution and macrophage response have not been studied. Molecular characterization of IBV strains circulating in Canada has been done previously and it appears that there is a difference in distribution of IBV genotypes according to the geographic area within Canada [
<xref ref-type="bibr" rid="CR33">33</xref>
,
<xref ref-type="bibr" rid="CR37">37</xref>
,
<xref ref-type="bibr" rid="CR50">50</xref>
,
<xref ref-type="bibr" rid="CR51">51</xref>
]. In Western Canada, mostly Mass and Conn IBV genotypes are common and in Eastern Canada, in addition to Mass and Conn genotypes nepharopathogenic strains, 4/91, CA1737 and DMV genotypes are common among poultry flocks.</p>
<p id="Par19">It appears that host responses, characterized by respiratory macrophages following IBV infection is dependent on the IBV strain. For example, it has been shown that M41 but not Australian T strain increases macrophages in respiratory tract lavage [
<xref ref-type="bibr" rid="CR64">64</xref>
]. Previously, it has also been shown that IBV ConnA5968 strain elicit higher macrophage response in tracheal mucosa when compared IBV M41 strain [
<xref ref-type="bibr" rid="CR65">65</xref>
]. Although Mass IBV (M41) strain is known to increase macrophage responses in lungs and trachea [
<xref ref-type="bibr" rid="CR65">65</xref>
], it is unknown whether variants of Mass IBV are capable of eliciting differential macrophage response in the respiratory mucosa. The current study shows that minor genetic differences in Mass IBV variants do not lead to differences in macrophage responses in the respiratory tract although each of the tested IBV isolates were able to elicit significant macrophage responses in these tissues. IBV M41 and ConnA5968 strains have been shown to replicate in macrophages in vivo and in vitro [
<xref ref-type="bibr" rid="CR65">65</xref>
<xref ref-type="bibr" rid="CR67">67</xref>
]. The IBV Beaudette strain is able to infect the HD11 macrophage cell line [
<xref ref-type="bibr" rid="CR68">68</xref>
]. Although we did not observe a difference in macrophage infection in vivo by both Mass variant IBV isolates used in our experiments, it will be important to establish if these field IBV strains also target macrophages located in the respiratory tract for its replication. We observed a higher quantity of IBV antigens in 15SK-02 IBV infected lungs when compared to 15AB-01 infected lungs and that difference was not reflected in the number of IBV infected macrophages. This discrepancy may be due to the fact that IBV infects cells other than macrophages. Previously, we observed that IBV infection in macrophage of the respiratory tract was about 3–4% [
<xref ref-type="bibr" rid="CR65">65</xref>
].</p>
<p id="Par20">In spite of the novel aspect of our work comparing two Mass type IBV isolates with minor genetic variations for pathogenicity and host responses, there were some limitations to this study. First, we did not include a control group infected with the standard M41 IBV strain in our experiment and this was due to the limited infrastructure availability at our animal facility. However, in a separate study, we investigated the M41 IBV induced macrophage recruitment to respiratory tract [
<xref ref-type="bibr" rid="CR65">65</xref>
]. Second, for the quantification of IBV, we employed real-time PCR technique targeting the most conserved N gene of IBV [
<xref ref-type="bibr" rid="CR69">69</xref>
<xref ref-type="bibr" rid="CR71">71</xref>
]. Since qPCR quantify both live and dead virus [
<xref ref-type="bibr" rid="CR72">72</xref>
], our IBV genome load data do not reflect IBV replication. Alternatively, we would have determined viral titers using 9–11 day old SPF embryonated chicken eggs and expressed as EID
<sub>50</sub>
[
<xref ref-type="bibr" rid="CR73">73</xref>
]. Although this later method could be employed to determine IBV infectivity in our swabs and tissues, the use of this technique was precluded for quantifying IBV titer due to the requirement of large numbre of eggs.</p>
</sec>
<sec id="Sec9">
<title>Conclusion</title>
<p id="Par21">In conclusion, Mass IBV variants recovered from layer flocks had identical nucleotide sequence at ORFs 3a, 3b, E, M, 5a and 5b. The remainder of the ORFs showed 99.9% nucleotide similarity. Although, the genetic changes were minor, the pathogenicity in vivo of the Mass IBV variant originating from AB was different to the one that originated in SK, whereby the AB variant caused significantly higher tracheal lesions and replicated to a lower titer in the lung and lower genome loads in cecal tonsils. Additionally, both Mass IBV variants elicited host responses characterized by significant macrophage recruitment to the respiratory tract and there was evidence that both Mass IBV variants replicated within macrophages in the respiratory tract. Studies are underway to evaluate the pathogenicity of these two Mass type IBV isolates in laying hens.</p>
</sec>
<sec id="Sec10">
<title>Methods</title>
<sec id="Sec11">
<title>Animals</title>
<p id="Par22">Day old white leghorn (SPF) layer chickens (White Leghorn) were obtained from the Canadian Food Inspection Agency (CFIA), Ottawa, ON, Canada and housed in high containment poultry isolators at the Prion- Virology Animal Facility of the University of Calgary’s Foothills Campus with ad libitum access to feed and water.</p>
</sec>
<sec id="Sec12">
<title>Virus</title>
<p id="Par23">Mass IBV isolates originating from layer flocks in AB (15AB-01) and SK (15SK-02) were used in the study. Viral titers were determined using 9-day old SPF embryonated chicken eggs and expressed as 50% embryo infectious dose (EID
<sub>50</sub>
) according to the method described by Reed and Muench [
<xref ref-type="bibr" rid="CR73">73</xref>
].</p>
</sec>
<sec id="Sec13">
<title>Experimental design</title>
<sec id="Sec14">
<title>Whole genome sequencing of mass IBV isolates that originated from Western Canada</title>
<p id="Par24">The Mass-type IBV isolates from the current study were sequentially passaged in 9 to 11-day old embryonated eggs and allantoic fluid was harvested and concentrated using ultracentrifugation at 50,000 g for 3 h. The resulting pellet was fractionated using the Optiprep™ kit (Millipore-Sigma, Missouri, USA). Ribonucleic acid (RNA) was extracted from the sixth fraction and allantoic fluid directly. cDNA was synthesized, and quantitative polymerase chain reaction (PCR) assay was performed to quantify the IBV genome load.</p>
</sec>
<sec id="Sec15">
<title>Infection of chickens with mass IBV isolates originated from Western Canada</title>
<p id="Par25">Two groups of 6 day-old unsexed-SPF chickens (
<italic>n</italic>
 = 7 per group) were infected with 50 μL inoculum containing 2.75× 10
<sup>4</sup>
PFUs of IBV isolates 15AB-01 and 15SK-02 intra-tracheally under isoflurane anesthesia. Another 5 of 6-day old SPF chickens were kept as uninfected controls. The chickens were weighed at 0, 2, 3, 4, 10, 12 and 14 dpi. At 2, 3, 10 and 12 dpi, oropharyngeal and cloacal swabs were obtained using Puritan® Unitranz-RT transport system (Puritan Medical Products LLC, Maine, USA) and stored at − 80 °C until further processing. At 4 dpi, 4 chickens from each infected and 3 chickens from the uninfected group were euthanized and trachea, lung, kidneys, cecal tonsils and reproductive tract samples were collected into RNASave® (Biological Industries, Beit Haemek, Israel) and stored at -20 °C. In addition, a portion of trachea, lung, kidneys, cecal tonsils and reproductive tract samples were collected into 10% neutral buffered formalin for histopathological examination and in Optimum Cutting Medium (OCT, Leica Biosystems, Wetzlar, Germany) for immunostaining. The remainder of chickens were continued to be observed and tissue samples were collected in RNASave® for genome load quantification at 14 dpi.</p>
<p id="Par26">For the sampling of tissues, the chickens were euthanized using overdose of isoflurane anesthesia followed by cervical dislocation.</p>
</sec>
</sec>
<sec id="Sec16">
<title>Techniques</title>
<sec id="Sec17">
<title>Sample preparation for whole genome sequencing</title>
<p id="Par27">The virus fractions, which gave the highest genome load or the lowest Ct were selected from both isolates and cDNA concentration was determined using the Qubit 4 Fluorometer (Thermo Fisher Scientific, MA, USA). A total of 10–12 ng of cDNA from each isolate was sent for whole genome sequencing to the Swine and Poultry Infectious Diseases Research Center (CRIPA), University of Montreal, Canada where library preparation and sequencing performed on Illumina® Miseq (San Diego, California, United States) next generation sequencing (NGS) platform generating 150 bp paired-end reads.</p>
</sec>
<sec id="Sec18">
<title>Histology</title>
<p id="Par28">Formalin-fixed terminal tissue samples were processed at the Diagnostic Services Unit of the University of Calgary’s Faculty of Veterinary Medicine including staining of sections with stained hematoxylin and eosin (H&E). Sections were examined under the light microscope and photomicrographs were taken under 40x magnification. The histological changes observed in the trachea [
<xref ref-type="bibr" rid="CR8">8</xref>
] and lungs [
<xref ref-type="bibr" rid="CR69">69</xref>
] were scored as described previously.</p>
</sec>
<sec id="Sec19">
<title>RNA extraction and cDNA synthesis</title>
<p id="Par29">Tracheal and cloacal swab samples, which were in the transport media were vortexed and medium was transferred to 1.5 mL centrifuge tubes and centrifuged at 1000 g for 20 mins at − 4 °C. A total of 250 μL of supernatant was used for RNA extraction using the Trizol LS® reagent (Ambion, Invitrogen Canada Inc., Burlington, ON, Canada) following manufacturer’s instructions. Approximately 50 mg piece of tissue preserved in RNA Save® was homogenized in 1 mL of Trizol® reagent (Ambion, Invitrogen Canada Inc., Burlington, ON, Canada) using a Pro200 Power homogenizer (Diamed, Mississauga, ON, Canada) on ice and RNA was extracted following the manufacturer’s protocol. The RNA pellet was re-suspended in 20 μL of RNase-free water and quantified using the Nanodrop 1000 spectrophotometer at 260 nm wavelength (Thermo Scientific, Wilmington, DE, USA). A total of 2000 ng of RNA was transcribed using 10X RT random primers (High Capacity cDNA Reverse Transcription Kit, Invitrogen Life Technologies, Carlsbad, CA, USA) as has been instructed by the manufacturer.</p>
</sec>
<sec id="Sec20">
<title>IBV genome load quantification</title>
<p id="Par30">The IBV genome load quantification was carried out following the products and protocol as described previously [
<xref ref-type="bibr" rid="CR65">65</xref>
]. Briefly, real-time PCR method was used for IBV genome load quantification using forward primer, IBV N Fw: 5’GACGGAGGACCTGATGGTAA3’ and reverse primer, IBV-N Re: 5’CCCTTCTTCTGCTGATCCTG3’. Thermal cycling conditions were 95 °C for 20 s; 40 cycles of amplification/extension at 95 °C for 3 s, and 60 °C for 30 s followed by a melting curve analysis was done between 65 °C to 95 °C with an increment of 0.5 °C at every 5 s. Fluorescent acquisition was done at 60 °C for 30 s.</p>
</sec>
<sec id="Sec21">
<title>Immunofluorescent assay</title>
<p id="Par31">Frozen tissues in OCT blocks were sectioned at 5 μm thickness using a cryotome (Leica Biosystems, Richmond, Illinois, USA) and adhered on to positively charged slides. The sections were air dried on the bench for 20 min, fixed by immersing in ice-cold acetone for 5 min and stained for macrophage and IBV antigens sequentially as has been described previously [
<xref ref-type="bibr" rid="CR65">65</xref>
]. Briefly, after protein blocking with 2.5% horse serum, the cryosections were incubated for 30 min with mouse monoclonal anti-chicken macrophage (KUL01) antibodies (Southern Biotech, Birmingham, Alabama, USA) (1:200 in PBS containing 2.5% horse serum). The staining specificity of KUL01 anti-chicken macrophage antibody was confirmed previously using an appropriate isotype control (Additional file
<xref rid="MOESM1" ref-type="media">1</xref>
: Figure 1). The secondary antibody, DyLight® 550 conjugated goat anti-mouse IgG (H + L) (Vector Laboratories, Inc., Burlingame, California, USA) (1:500 in PBS containing 2.5% horse serum) was used. Before being stained for IBV antigen using anti-IBV rabbit polyclonal serum (Federal Research Institute for Animal Health, Greifswald-Insel Riems, Germany; 1:3000 diluted in 2.5% horse serum), the sections were blocked again with 2.5% of horse serum. For subsequent staining, Vectaflour RTU Antibody kit containing Dylight 488 conjugate (DK-1488, Vector Laboratories, Burlingame, California, USA). The sections were mounted on Vectashield® antifade mounting medium with 4′,6-Diamidine-2′-phenylindole dihydrochloride (DAPI) nuclear stain (Vector Laboratories, Burlingame, California, USA).</p>
</sec>
</sec>
<sec id="Sec22">
<title>Data analyses</title>
<sec id="Sec23">
<title>Whole genome sequencing data analysis</title>
<p id="Par32">All reads were assembled to get a single contig. The resulting full genome sequences were aligned, and identity matrix was performed with 43 published sequences of IBV reference serotypes retrieved from the National Center for Biotechnology Information (NCBI) GenBank database using the BLAST® research tool and the Custal Omega 1.2.3 package with automated settings (Geneious® 10.1.3). The phylogenetic tree inferred based on the full-length IBV genomes using Randomized Axelerated Maximum Likelihood (RAxML 8.2.7) method with GTR GAMMA nucleotide substitution model with 1000 bootstrap iterations (Geneious software, version 10.1.3, Biomatters Ltd., Auckland, New Zealand). The resulting proportional tree of the two Mass IBV isolates of this study with 43 reference sequences (Additional file 
<xref rid="MOESM2" ref-type="media">2</xref>
: Table S1) is shown with the bootstrap support. Nucleotide sequences of the two Mass IBV isolates, 15AB-01 and 15SK-02 were deposited in the NCBI GenBank public domain under the accession numbers MH539771 and MH539772, respectively. ORFs of the two new sequences were determined using the NCBI ORF finder (
<ext-link ext-link-type="uri" xlink:href="https://www.ncbi.nlm.nih.gov/orffinder/">https://www.ncbi.nlm.nih.gov/orffinder/</ext-link>
) and the genome annotation was carried out using Geneious software, version 10.1.3 (Biomatters Ltd., Auckland, New Zealand).</p>
</sec>
<sec id="Sec24">
<title>IBV genome load quantification</title>
<p id="Par33">IBV genome copies per 200 ng of cDNA were calculated using a standard curve based on a serial dilution of plasmids as described previously [
<xref ref-type="bibr" rid="CR69">69</xref>
].</p>
</sec>
<sec id="Sec25">
<title>Quantification of the number of macrophages, IBV antigen and IBV positive macrophages</title>
<p id="Par34">For the quantification of macrophages and the IBV antigen in the trachea and lungs, five areas with the highest DyLight® 550 (macrophages) and DyLight® 488 (IBV antigen) fluorescent signals and corresponding nuclear stained (DAPI) areas were captured under 40x magnification using an epifluorescent microscope (Olympus IX51, Center Valley, Pensylvania, USA). The obtained images were transferred to the Image-J® software (National Institute of Health, Bethesda, Maryland, USA) and the areas of each micrograph covered with DyLight® 550 (macrophages), DyLight® 488 (IBV) signals, combined DyLight® 550 and DyLight® 488 signals (IBV positive macrophages) and the combined DyLight® 550 and DyLight® 488 signals with DAPI (total area of the section) were quantified. The areas covered with macrophage and IBV antigens and the combined signals were expressed as a percentage of the total area covered by the section.</p>
</sec>
<sec id="Sec26">
<title>Statistical analyses</title>
<p id="Par35">Two-way analysis of variance (ANOVA) followed by Benferroni post-hoc test was used to determine the significant time points of body weight among the infected and control groups. Tracheal and lung histopathological lesion score data was analyzed using Kruskal-Wallis test followed by Dunn’s multiple comparison test. The genome load data was analyzed by the two-way ANOVA followed by Bonferroni post-test. One-way ANOVA test followed by Tukey’s posttest was used to determine the significance of immunofluorescence data. Grubbs’ outlier test was performed in order to identify outliers before the data was analyzed. All the statistical procedures were performed using in-built statistical analysis feature of the GraphPad™ Prism 5 (GraphPad Software, La Jolla, California, USA). Differences were considered significant at
<italic>p</italic>
 < 0.05. The significance on graphs are labelled as the following: *
<italic>p</italic>
 < 0.05, **
<italic>p</italic>
 < 0.01, ***
<italic>p</italic>
 < 0.001, and ****
<italic>p</italic>
 < 0.0001.</p>
</sec>
</sec>
</sec>
<sec sec-type="supplementary-material">
<title>Additional files</title>
<sec id="Sec27">
<p>
<supplementary-material content-type="local-data" id="MOESM1">
<media xlink:href="12917_2018_1720_MOESM1_ESM.docx">
<label>Additional file 1:</label>
<caption>
<p>
<bold>Figure S1.</bold>
Specificity of avian macrophage staining. The power point slide contain lung sections stained with anti-chicken macrophage antibody (KUL-01) and isotype control. The sections were also stained with nuclear staining (DAPI). (DOCX 271 kb).</p>
</caption>
</media>
</supplementary-material>
<supplementary-material content-type="local-data" id="MOESM2">
<media xlink:href="12917_2018_1720_MOESM2_ESM.docx">
<label>Additional file 2:</label>
<caption>
<p>
<bold>Table S1.</bold>
List of reference IBV sequences used in this study. This table contain names and accession numbers of IBV whole genome reference sequences retrieved from the GenBank repository. (DOCX 15 kb).</p>
</caption>
</media>
</supplementary-material>
</p>
</sec>
</sec>
</body>
<back>
<ack>
<title>Acknowledgements</title>
<p>We acknowledge the staff involved in field poultry veterinary services in SK and AB, Canada. We acknowledge the staff of Prion-Virology Animal Facility, University of Calgary for routine monitoring of experimental animals.</p>
<sec id="FPar1">
<title>Funding</title>
<p id="Par36">This research including graduate student stipend was funded by the SK Agricultural Development Fund and SK Egg Producers (Project number: 20140185). Graduate student, AA was also supported partly by University of Calgary’s Eyes-High International Doctoral Scholarship and Bridge Funding from the Department of Ecosystem and Public Health. These funding bodies were not involved in the design of the study, collection, analysis, interpretation of data and in writing the manuscript.</p>
</sec>
<sec id="FPar2">
<title>Availability of data and materials</title>
<p id="Par37">Nucleotide sequences of the two Mass IBV isolates, 15AB-01 and 15SK-02 is available in the NCBI GenBank public domain under the accession numbers, MH539771 and MH539772, respectively. The remainder of datasets used during the current study are available from the corresponding author on request.</p>
</sec>
</ack>
<notes notes-type="author-contribution">
<title>Authors’ contributions</title>
<p>Conceptualization, funding acquisition and project administration was done by SG and MFA. Investigations, data collection and formal analysis were undertaken by AA, UDSS and MSA. SP, FM and Saskatchewan Poultry Extension (Tennille Knezacek, Jenny Fricke and Stephanie Derbawka) helped in sample collection leading to the isolation of Mass IBV isolates. Original draft was prepared by AA. Review of the manuscript including editing and student supervision and mentoring were done by SG, SCC, FvdM and MFA. All authors read and approved the final manuscript.</p>
</notes>
<notes notes-type="COI-statement">
<sec id="FPar3">
<title>Authors’ information</title>
<p>AA: PhD student, University of Calgary.</p>
<p>UDSS: MSc student, University of Calgary.</p>
<p>MSA: PhD student, University of Calgary.</p>
<p>SP: Research Technician-Poultry, University of Saskatchewan.</p>
<p>FM: Veterinarian, Marshall Swine and Poultry Health Services.</p>
<p>SCC: Professor and Head, Ecosystem and Public Health, University of Calgary.</p>
<p>FvdM: Associate Professor (Global Health), University of Calgary.</p>
<p>SG: Professor and Head, Department of Veterinary Pathology, University of Saskatchewan.</p>
<p>MFA: Associate Professor (Virology), University of Calgary.</p>
</sec>
<sec id="FPar4">
<title>Ethics approval</title>
<p>The HSACC of the University of Calgary approved the use of SPF chickens used in all our experimental procedures (Protocol number AC15–0140).</p>
</sec>
<sec id="FPar5">
<title>Consent for publication</title>
<p>Not applicable</p>
</sec>
<sec id="FPar6">
<title>Competing interests</title>
<p>The authors declare that they have no competing interests.</p>
</sec>
<sec id="FPar7">
<title>Publisher’s Note</title>
<p>Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.</p>
</sec>
</notes>
<ref-list id="Bib1">
<title>References</title>
<ref id="CR1">
<label>1.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Cavanagh</surname>
<given-names>D</given-names>
</name>
</person-group>
<article-title>Coronaviruses in poultry and other birds</article-title>
<source>Avian Pathology</source>
<year>2005</year>
<volume>34</volume>
<issue>6</issue>
<fpage>439</fpage>
<lpage>448</lpage>
<pub-id pub-id-type="pmid">16537157</pub-id>
</element-citation>
</ref>
<ref id="CR2">
<label>2.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Cavanagh</surname>
<given-names>D</given-names>
</name>
</person-group>
<article-title>Coronavirus avian infectious bronchitis virus</article-title>
<source>Vet Res</source>
<year>2007</year>
<volume>38</volume>
<issue>2</issue>
<fpage>281</fpage>
<lpage>297</lpage>
<pub-id pub-id-type="pmid">17296157</pub-id>
</element-citation>
</ref>
<ref id="CR3">
<label>3.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Cumming</surname>
<given-names>RB</given-names>
</name>
</person-group>
<article-title>The control of avian infectious bronchitis/nephrosis in Australia</article-title>
<source>Aust Vet J</source>
<year>1969</year>
<volume>45</volume>
<issue>4</issue>
<fpage>200</fpage>
<lpage>203</lpage>
<pub-id pub-id-type="pmid">5814107</pub-id>
</element-citation>
</ref>
<ref id="CR4">
<label>4.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Kingham</surname>
<given-names>BF</given-names>
</name>
<name>
<surname>Keeler</surname>
<given-names>CL</given-names>
</name>
<name>
<surname>Nix</surname>
<given-names>WA</given-names>
</name>
<name>
<surname>Ladman</surname>
<given-names>BS</given-names>
</name>
<name>
<surname>Gelb</surname>
<given-names>J</given-names>
</name>
</person-group>
<article-title>Identification of avian infectious bronchitis virus by direct automated cycle sequencing of the S-1 gene</article-title>
<source>Avian Dis</source>
<year>2000</year>
<volume>44</volume>
<issue>2</issue>
<fpage>325</fpage>
<lpage>335</lpage>
<pub-id pub-id-type="pmid">10879913</pub-id>
</element-citation>
</ref>
<ref id="CR5">
<label>5.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Song</surname>
<given-names>C-S</given-names>
</name>
<name>
<surname>Lee</surname>
<given-names>Y-J</given-names>
</name>
<name>
<surname>Lee</surname>
<given-names>C-W</given-names>
</name>
<name>
<surname>Sung</surname>
<given-names>H-W</given-names>
</name>
<name>
<surname>Kim</surname>
<given-names>J-H</given-names>
</name>
<name>
<surname>Mo</surname>
<given-names>I-P</given-names>
</name>
<name>
<surname>Izumiya</surname>
<given-names>Y</given-names>
</name>
<name>
<surname>Jang</surname>
<given-names>H-K</given-names>
</name>
<name>
<surname>Mikami</surname>
<given-names>T</given-names>
</name>
</person-group>
<article-title>Induction of protective immunity in chickens vaccinated with infectious bronchitis virus S1 glycoprotein expressed by a recombinant baculovirus</article-title>
<source>J Gen Virol</source>
<year>1998</year>
<volume>79</volume>
<issue>4</issue>
<fpage>719</fpage>
<lpage>723</lpage>
<pub-id pub-id-type="pmid">9568966</pub-id>
</element-citation>
</ref>
<ref id="CR6">
<label>6.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Liu</surname>
<given-names>S</given-names>
</name>
<name>
<surname>Chen</surname>
<given-names>J</given-names>
</name>
<name>
<surname>Han</surname>
<given-names>Z</given-names>
</name>
<name>
<surname>Zhang</surname>
<given-names>Q</given-names>
</name>
<name>
<surname>Shao</surname>
<given-names>Y</given-names>
</name>
<name>
<surname>Kong</surname>
<given-names>X</given-names>
</name>
<name>
<surname>Tong</surname>
<given-names>G</given-names>
</name>
</person-group>
<article-title>Infectious bronchitis virus: S1 gene characteristics of vaccines used in China and efficacy of vaccination against heterologous strains from China</article-title>
<source>Avian Pathology</source>
<year>2006</year>
<volume>35</volume>
<issue>5</issue>
<fpage>394</fpage>
<lpage>399</lpage>
<pub-id pub-id-type="pmid">16990149</pub-id>
</element-citation>
</ref>
<ref id="CR7">
<label>7.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Reddy</surname>
<given-names>VRAP</given-names>
</name>
<name>
<surname>Theuns</surname>
<given-names>S</given-names>
</name>
<name>
<surname>Roukaerts</surname>
<given-names>IDM</given-names>
</name>
<name>
<surname>Zeller</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Matthijnssens</surname>
<given-names>J</given-names>
</name>
<name>
<surname>Nauwynck</surname>
<given-names>HJ</given-names>
</name>
</person-group>
<article-title>Genetic characterization of the Belgian Nephropathogenic infectious bronchitis virus (NIBV) reference strain B1648</article-title>
<source>Viruses</source>
<year>2015</year>
<volume>7</volume>
<issue>8</issue>
<fpage>4488</fpage>
<lpage>4506</lpage>
<pub-id pub-id-type="pmid">26262637</pub-id>
</element-citation>
</ref>
<ref id="CR8">
<label>8.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Grgic</surname>
<given-names>H</given-names>
</name>
<name>
<surname>Hunter</surname>
<given-names>DB</given-names>
</name>
<name>
<surname>Hunton</surname>
<given-names>P</given-names>
</name>
<name>
<surname>Nagy</surname>
<given-names>E</given-names>
</name>
</person-group>
<article-title>Pathogenicity of infectious bronchitis virus isolates from Ontario chickens</article-title>
<source>Can J Vet Res</source>
<year>2008</year>
<volume>72</volume>
<issue>5</issue>
<fpage>403</fpage>
<lpage>410</lpage>
<pub-id pub-id-type="pmid">19086372</pub-id>
</element-citation>
</ref>
<ref id="CR9">
<label>9.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Boroomand</surname>
<given-names>Z</given-names>
</name>
<name>
<surname>Asasi</surname>
<given-names>K</given-names>
</name>
<name>
<surname>Mohammadi</surname>
<given-names>A</given-names>
</name>
</person-group>
<article-title>Pathogenesis and tissue distribution of avian infectious bronchitis virus isolate IRFIBV32 (793/B serotype) in experimentally infected broiler chickens</article-title>
<source>Sci World J</source>
<year>2012</year>
<volume>2012</volume>
<fpage>402537</fpage>
</element-citation>
</ref>
<ref id="CR10">
<label>10.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Franca</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Woolcock</surname>
<given-names>PR</given-names>
</name>
<name>
<surname>Yu</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Jackwood</surname>
<given-names>MW</given-names>
</name>
<name>
<surname>Shivaprasad</surname>
<given-names>HL</given-names>
</name>
</person-group>
<article-title>Nephritis associated with infectious bronchitis virus Cal99 variant in game chickens</article-title>
<source>Avian Dis</source>
<year>2011</year>
<volume>55</volume>
<issue>3</issue>
<fpage>422</fpage>
<lpage>428</lpage>
<pub-id pub-id-type="pmid">22017040</pub-id>
</element-citation>
</ref>
<ref id="CR11">
<label>11.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Ambali</surname>
<given-names>AG</given-names>
</name>
<name>
<surname>Jones</surname>
<given-names>RC</given-names>
</name>
</person-group>
<article-title>Early pathogenesis in chicks of infection with an Enterotropic strain of infectious bronchitis virus</article-title>
<source>Avian Dis</source>
<year>1990</year>
<volume>34</volume>
<issue>4</issue>
<fpage>809</fpage>
<lpage>817</lpage>
<pub-id pub-id-type="pmid">2177973</pub-id>
</element-citation>
</ref>
<ref id="CR12">
<label>12.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>El-Houadfi</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Jones</surname>
<given-names>RC</given-names>
</name>
<name>
<surname>Cook</surname>
<given-names>JKA</given-names>
</name>
<name>
<surname>Ambali</surname>
<given-names>AG</given-names>
</name>
</person-group>
<article-title>The isolation and characterisation of six avian infectious bronchitis viruses isolated in Morocco</article-title>
<source>Avian Pathology</source>
<year>1986</year>
<volume>15</volume>
<issue>1</issue>
<fpage>93</fpage>
<lpage>105</lpage>
<pub-id pub-id-type="pmid">18766508</pub-id>
</element-citation>
</ref>
<ref id="CR13">
<label>13.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Sevoian</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Levine</surname>
<given-names>P</given-names>
</name>
</person-group>
<article-title>Effects of infectious bronchitis on the reproductive tracts, egg production, and egg quality of laying chickens</article-title>
<source>Avian Dis</source>
<year>1957</year>
<volume>1</volume>
<issue>2</issue>
<fpage>136</fpage>
<lpage>164</lpage>
</element-citation>
</ref>
<ref id="CR14">
<label>14.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>MacDonald</surname>
<given-names>JW</given-names>
</name>
<name>
<surname>McMartin</surname>
<given-names>DA</given-names>
</name>
</person-group>
<article-title>Observations on the effects of the h52 and h120 vaccine strains of infectious bronchitis virus in the domestic fowl</article-title>
<source>Avian Pathology</source>
<year>1976</year>
<volume>5</volume>
<issue>3</issue>
<fpage>157</fpage>
<lpage>173</lpage>
<pub-id pub-id-type="pmid">18777342</pub-id>
</element-citation>
</ref>
<ref id="CR15">
<label>15.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Raj</surname>
<given-names>GD</given-names>
</name>
<name>
<surname>Jones</surname>
<given-names>RC</given-names>
</name>
</person-group>
<article-title>Infectious bronchitis virus: Immunopathogenesis of infection in the chicken</article-title>
<source>Avian Pathology</source>
<year>1997</year>
<volume>26</volume>
<fpage>677</fpage>
<lpage>706</lpage>
<pub-id pub-id-type="pmid">18483939</pub-id>
</element-citation>
</ref>
<ref id="CR16">
<label>16.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Cook</surname>
<given-names>JK</given-names>
</name>
<name>
<surname>Jackwood</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Jones</surname>
<given-names>RC</given-names>
</name>
</person-group>
<article-title>The long view: 40 years of infectious bronchitis research</article-title>
<source>Avian Pathology</source>
<year>2012</year>
<volume>41</volume>
<issue>3</issue>
<fpage>239</fpage>
<lpage>250</lpage>
<pub-id pub-id-type="pmid">22702451</pub-id>
</element-citation>
</ref>
<ref id="CR17">
<label>17.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Crinion</surname>
<given-names>RAP</given-names>
</name>
<name>
<surname>Ball</surname>
<given-names>RA</given-names>
</name>
<name>
<surname>Hofstad</surname>
<given-names>MS</given-names>
</name>
</person-group>
<article-title>Abnormalities in laying chickens following exposure to infectious bronchitis virus at one day old</article-title>
<source>Avian Dis</source>
<year>1971</year>
<volume>15</volume>
<issue>1</issue>
<fpage>42</fpage>
<lpage>48</lpage>
<pub-id pub-id-type="pmid">5547755</pub-id>
</element-citation>
</ref>
<ref id="CR18">
<label>18.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Benyeda</surname>
<given-names>Z</given-names>
</name>
<name>
<surname>Mato</surname>
<given-names>T</given-names>
</name>
<name>
<surname>Suveges</surname>
<given-names>T</given-names>
</name>
<name>
<surname>Szabo</surname>
<given-names>E</given-names>
</name>
<name>
<surname>Kardi</surname>
<given-names>V</given-names>
</name>
<name>
<surname>Abonyi-Toth</surname>
<given-names>Z</given-names>
</name>
<name>
<surname>Rusvai</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Palya</surname>
<given-names>V</given-names>
</name>
</person-group>
<article-title>Comparison of the pathogenicity of QX-like, M41 and 793/B infectious bronchitis strains from different pathological conditions</article-title>
<source>Avian Pathology</source>
<year>2009</year>
<volume>38</volume>
<issue>6</issue>
<fpage>449</fpage>
<lpage>456</lpage>
<pub-id pub-id-type="pmid">19937534</pub-id>
</element-citation>
</ref>
<ref id="CR19">
<label>19.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Boltz</surname>
<given-names>DA</given-names>
</name>
<name>
<surname>Nakai</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Bahra</surname>
<given-names>JM</given-names>
</name>
</person-group>
<article-title>Avian infectious bronchitis virus: a possible cause of reduced fertility in the rooster</article-title>
<source>Avian Dis</source>
<year>2004</year>
<volume>48</volume>
<issue>4</issue>
<fpage>909</fpage>
<lpage>915</lpage>
<pub-id pub-id-type="pmid">15666874</pub-id>
</element-citation>
</ref>
<ref id="CR20">
<label>20.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Liu</surname>
<given-names>S</given-names>
</name>
<name>
<surname>Zhang</surname>
<given-names>X</given-names>
</name>
<name>
<surname>Wang</surname>
<given-names>Y</given-names>
</name>
<name>
<surname>Li</surname>
<given-names>C</given-names>
</name>
<name>
<surname>Han</surname>
<given-names>Z</given-names>
</name>
<name>
<surname>Shao</surname>
<given-names>Y</given-names>
</name>
<name>
<surname>Li</surname>
<given-names>H</given-names>
</name>
<name>
<surname>Kong</surname>
<given-names>X</given-names>
</name>
</person-group>
<article-title>Molecular characterization and pathogenicity of infectious bronchitis coronaviruses: complicated evolution and epidemiology in China caused by Cocirculation of multiple types of infectious bronchitis coronaviruses</article-title>
<source>Intervirology</source>
<year>2009</year>
<volume>52</volume>
<issue>4</issue>
<fpage>223</fpage>
<lpage>234</lpage>
<pub-id pub-id-type="pmid">19590226</pub-id>
</element-citation>
</ref>
<ref id="CR21">
<label>21.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Alvarado</surname>
<given-names>IR</given-names>
</name>
<name>
<surname>Villegas</surname>
<given-names>P</given-names>
</name>
<name>
<surname>Mossos</surname>
<given-names>N</given-names>
</name>
<name>
<surname>Jackwood</surname>
<given-names>MW</given-names>
</name>
</person-group>
<article-title>Molecular characterization of avian infectious bronchitis virus strains isolated in Colombia during 2003</article-title>
<source>Avian Dis</source>
<year>2005</year>
<volume>49</volume>
<issue>4</issue>
<fpage>494</fpage>
<lpage>499</lpage>
<pub-id pub-id-type="pmid">16404989</pub-id>
</element-citation>
</ref>
<ref id="CR22">
<label>22.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Kusters</surname>
<given-names>JG</given-names>
</name>
<name>
<surname>Niesters</surname>
<given-names>HGM</given-names>
</name>
<name>
<surname>Bleumink-Pluym</surname>
<given-names>NMC</given-names>
</name>
<name>
<surname>Davelaar</surname>
<given-names>FG</given-names>
</name>
<name>
<surname>Horzinek</surname>
<given-names>MC</given-names>
</name>
<name>
<surname>Van der Zeijst</surname>
<given-names>BAM</given-names>
</name>
</person-group>
<article-title>Molecular epidemiology of infectious bronchitis virus in the Netherlands</article-title>
<source>J Gen Virol</source>
<year>1987</year>
<volume>68</volume>
<issue>2</issue>
<fpage>343</fpage>
<lpage>352</lpage>
<pub-id pub-id-type="pmid">3029279</pub-id>
</element-citation>
</ref>
<ref id="CR23">
<label>23.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Lee</surname>
<given-names>C-W</given-names>
</name>
<name>
<surname>Jackwood</surname>
<given-names>MW</given-names>
</name>
</person-group>
<article-title>Evidence of genetic diversity generated by recombination among avian coronavirus IBV</article-title>
<source>Arch Virol</source>
<year>2000</year>
<volume>145</volume>
<issue>10</issue>
<fpage>2135</fpage>
<lpage>2148</lpage>
<pub-id pub-id-type="pmid">11087096</pub-id>
</element-citation>
</ref>
<ref id="CR24">
<label>24.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Kottier</surname>
<given-names>SA</given-names>
</name>
<name>
<surname>Cavanagh</surname>
<given-names>D</given-names>
</name>
<name>
<surname>Britton</surname>
<given-names>P</given-names>
</name>
</person-group>
<article-title>Experimental evidence of recombination in coronavirus infectious bronchitis virus</article-title>
<source>Virology</source>
<year>1995</year>
<volume>213</volume>
<issue>2</issue>
<fpage>569</fpage>
<lpage>580</lpage>
<pub-id pub-id-type="pmid">7491781</pub-id>
</element-citation>
</ref>
<ref id="CR25">
<label>25.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Wang</surname>
<given-names>L</given-names>
</name>
<name>
<surname>Xu</surname>
<given-names>Y</given-names>
</name>
<name>
<surname>Collisson</surname>
<given-names>EW</given-names>
</name>
</person-group>
<article-title>Experimental confirmation of recombination upstream of the S1 hypervariable region of infectious bronchitis virus</article-title>
<source>Virus Res</source>
<year>1997</year>
<volume>49</volume>
<issue>2</issue>
<fpage>139</fpage>
<lpage>145</lpage>
<pub-id pub-id-type="pmid">9213388</pub-id>
</element-citation>
</ref>
<ref id="CR26">
<label>26.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Mardani</surname>
<given-names>K</given-names>
</name>
<name>
<surname>Noormohammadi</surname>
<given-names>A</given-names>
</name>
<name>
<surname>Ignjatovic</surname>
<given-names>J</given-names>
</name>
<name>
<surname>Browning</surname>
<given-names>G</given-names>
</name>
</person-group>
<article-title>Naturally occurring recombination between distant strains of infectious bronchitis virus</article-title>
<source>Arch Virol</source>
<year>2010</year>
<volume>155</volume>
<issue>10</issue>
<fpage>1581</fpage>
<lpage>1586</lpage>
<pub-id pub-id-type="pmid">20574643</pub-id>
</element-citation>
</ref>
<ref id="CR27">
<label>27.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Jia</surname>
<given-names>W</given-names>
</name>
<name>
<surname>Karaca</surname>
<given-names>K</given-names>
</name>
<name>
<surname>Parrish</surname>
<given-names>CR</given-names>
</name>
<name>
<surname>Naqi</surname>
<given-names>SA</given-names>
</name>
</person-group>
<article-title>A novel variant of avian infectious bronchitis virus resulting from recombination among three different strains</article-title>
<source>Arch Virol</source>
<year>1995</year>
<volume>140</volume>
<issue>2</issue>
<fpage>259</fpage>
<lpage>271</lpage>
<pub-id pub-id-type="pmid">7710354</pub-id>
</element-citation>
</ref>
<ref id="CR28">
<label>28.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Montassier</surname>
<given-names>HJ</given-names>
</name>
</person-group>
<article-title>Molecular epidemiology and evolution of avian infectious bronchitis virus</article-title>
<source>Revista Brasileira de Ciência Avícola</source>
<year>2010</year>
<volume>12</volume>
<fpage>87</fpage>
<lpage>96</lpage>
</element-citation>
</ref>
<ref id="CR29">
<label>29.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Cavanagh</surname>
<given-names>D</given-names>
</name>
<name>
<surname>Davis</surname>
<given-names>PJ</given-names>
</name>
<name>
<surname>Darbyshire</surname>
<given-names>JH</given-names>
</name>
<name>
<surname>Peters</surname>
<given-names>RW</given-names>
</name>
</person-group>
<article-title>Coronavirus IBV: virus retaining spike glycopolypeptide S2 but not S1 is unable to induce virus-neutralizing or haemagglutination-inhibiting antibody, or induce chicken tracheal protection</article-title>
<source>J Gen Virol</source>
<year>1986</year>
<volume>67</volume>
<issue>7</issue>
<fpage>1435</fpage>
<lpage>1442</lpage>
<pub-id pub-id-type="pmid">3014053</pub-id>
</element-citation>
</ref>
<ref id="CR30">
<label>30.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Cavanagh</surname>
<given-names>D</given-names>
</name>
<name>
<surname>Davis</surname>
<given-names>PJ</given-names>
</name>
<name>
<surname>Cook</surname>
<given-names>JKA</given-names>
</name>
</person-group>
<article-title>Infectious bronchitis virus: evidence for recombination within the Massachusetts serotype</article-title>
<source>Avian Pathology</source>
<year>1992</year>
<volume>21</volume>
<issue>3</issue>
<fpage>401</fpage>
<lpage>408</lpage>
<pub-id pub-id-type="pmid">18670955</pub-id>
</element-citation>
</ref>
<ref id="CR31">
<label>31.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Selim</surname>
<given-names>K</given-names>
</name>
<name>
<surname>Arafa</surname>
<given-names>AS</given-names>
</name>
<name>
<surname>Hussein</surname>
<given-names>HA</given-names>
</name>
<name>
<surname>El-Sanousi</surname>
<given-names>AA</given-names>
</name>
</person-group>
<article-title>Molecular characterization of infectious bronchitis viruses isolated from broiler and layer chicken farms in Egypt during 2012</article-title>
<source>International Journal of Veterinary Science and Medicine</source>
<year>2013</year>
<volume>1</volume>
<issue>2</issue>
<fpage>102</fpage>
<lpage>108</lpage>
</element-citation>
</ref>
<ref id="CR32">
<label>32.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Awad</surname>
<given-names>F</given-names>
</name>
<name>
<surname>Baylis</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Ganapathy</surname>
<given-names>K</given-names>
</name>
</person-group>
<article-title>Detection of variant infectious bronchitis viruses in broiler flocks in Libya</article-title>
<source>International Journal of Veterinary Science and Medicine</source>
<year>2014</year>
<volume>2</volume>
<issue>1</issue>
<fpage>78</fpage>
<lpage>82</lpage>
</element-citation>
</ref>
<ref id="CR33">
<label>33.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Martin</surname>
<given-names>EA</given-names>
</name>
<name>
<surname>Brash</surname>
<given-names>ML</given-names>
</name>
<name>
<surname>Hoyland</surname>
<given-names>SK</given-names>
</name>
<name>
<surname>Coventry</surname>
<given-names>JM</given-names>
</name>
<name>
<surname>Sandrock</surname>
<given-names>C</given-names>
</name>
<name>
<surname>Guerin</surname>
<given-names>MT</given-names>
</name>
<name>
<surname>Ojkic</surname>
<given-names>D</given-names>
</name>
</person-group>
<article-title>Genotyping of infectious bronchitis viruses identified in Canada between 2000 and 2013</article-title>
<source>Avian Pathology</source>
<year>2014</year>
<volume>43</volume>
<issue>3</issue>
<fpage>264</fpage>
<lpage>268</lpage>
<pub-id pub-id-type="pmid">24754612</pub-id>
</element-citation>
</ref>
<ref id="CR34">
<label>34.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Jackwood</surname>
<given-names>MW</given-names>
</name>
<name>
<surname>Lee</surname>
<given-names>DH</given-names>
</name>
</person-group>
<article-title>Different evolutionary trajectories of vaccine-controlled and non-controlled avian infectious bronchitis viruses in commercial poultry</article-title>
<source>PLoS One</source>
<year>2017</year>
<volume>12</volume>
<issue>5</issue>
<fpage>e0176709</fpage>
<pub-id pub-id-type="pmid">28472110</pub-id>
</element-citation>
</ref>
<ref id="CR35">
<label>35.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Hong</surname>
<given-names>S-M</given-names>
</name>
<name>
<surname>Kwon</surname>
<given-names>H-J</given-names>
</name>
<name>
<surname>Kim</surname>
<given-names>I-H</given-names>
</name>
<name>
<surname>Mo</surname>
<given-names>M-L</given-names>
</name>
<name>
<surname>Kim</surname>
<given-names>J-H</given-names>
</name>
</person-group>
<article-title>Comparative genomics of Korean infectious bronchitis viruses (IBVs) and an animal model to evaluate pathogenicity of IBVs to the reproductive organs</article-title>
<source>Viruses</source>
<year>2012</year>
<volume>4</volume>
<issue>11</issue>
<fpage>2670</fpage>
<pub-id pub-id-type="pmid">23202499</pub-id>
</element-citation>
</ref>
<ref id="CR36">
<label>36.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Marandino</surname>
<given-names>A</given-names>
</name>
<name>
<surname>Pereda</surname>
<given-names>A</given-names>
</name>
<name>
<surname>Tomas</surname>
<given-names>G</given-names>
</name>
<name>
<surname>Hernandez</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Iraola</surname>
<given-names>G</given-names>
</name>
<name>
<surname>Craig</surname>
<given-names>MI</given-names>
</name>
<name>
<surname>Hernandez</surname>
<given-names>D</given-names>
</name>
<name>
<surname>Banda</surname>
<given-names>A</given-names>
</name>
<name>
<surname>Villegas</surname>
<given-names>P</given-names>
</name>
<name>
<surname>Panzera</surname>
<given-names>Y</given-names>
</name>
<etal></etal>
</person-group>
<article-title>Phylodynamic analysis of avian infectious bronchitis virus in South America</article-title>
<source>The Journal of General Virology</source>
<year>2015</year>
<volume>96</volume>
<issue>Pt 6</issue>
<fpage>1340</fpage>
<lpage>1346</lpage>
<pub-id pub-id-type="pmid">25667323</pub-id>
</element-citation>
</ref>
<ref id="CR37">
<label>37.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Smati</surname>
<given-names>R</given-names>
</name>
<name>
<surname>Silim</surname>
<given-names>A</given-names>
</name>
<name>
<surname>Guertin</surname>
<given-names>C</given-names>
</name>
<name>
<surname>Henrichon</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Marandi</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Arella</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Merzouki</surname>
<given-names>A</given-names>
</name>
</person-group>
<article-title>Molecular characterization of three new avian infectious bronchitis virus (IBV) strains isolated in Quebec</article-title>
<source>Virus Genes</source>
<year>2002</year>
<volume>25</volume>
<issue>1</issue>
<fpage>85</fpage>
<lpage>93</lpage>
<pub-id pub-id-type="pmid">12206312</pub-id>
</element-citation>
</ref>
<ref id="CR38">
<label>38.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Feng</surname>
<given-names>K</given-names>
</name>
<name>
<surname>Wang</surname>
<given-names>F</given-names>
</name>
<name>
<surname>Xue</surname>
<given-names>Y</given-names>
</name>
<name>
<surname>Zhou</surname>
<given-names>Q</given-names>
</name>
<name>
<surname>Chen</surname>
<given-names>F</given-names>
</name>
<name>
<surname>Bi</surname>
<given-names>Y</given-names>
</name>
<name>
<surname>Xie</surname>
<given-names>Q</given-names>
</name>
</person-group>
<article-title>Epidemiology and characterization of avian infectious bronchitis virus strains circulating in southern China during the period from 2013-2015</article-title>
<source>Sci Rep</source>
<year>2017</year>
<volume>7</volume>
<issue>1</issue>
<fpage>6576</fpage>
<pub-id pub-id-type="pmid">28747730</pub-id>
</element-citation>
</ref>
<ref id="CR39">
<label>39.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Mo</surname>
<given-names>ML</given-names>
</name>
<name>
<surname>Li</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Huang</surname>
<given-names>BC</given-names>
</name>
<name>
<surname>Fan</surname>
<given-names>WS</given-names>
</name>
<name>
<surname>Wei</surname>
<given-names>P</given-names>
</name>
<name>
<surname>Wei</surname>
<given-names>TC</given-names>
</name>
<name>
<surname>Cheng</surname>
<given-names>QY</given-names>
</name>
<name>
<surname>Wei</surname>
<given-names>ZJ</given-names>
</name>
<name>
<surname>Lang</surname>
<given-names>YH</given-names>
</name>
</person-group>
<article-title>Molecular characterization of major structural protein genes of avian coronavirus infectious bronchitis virus isolates in southern China</article-title>
<source>Viruses</source>
<year>2013</year>
<volume>5</volume>
<issue>12</issue>
<fpage>3007</fpage>
<lpage>3020</lpage>
<pub-id pub-id-type="pmid">24304696</pub-id>
</element-citation>
</ref>
<ref id="CR40">
<label>40.</label>
<mixed-citation publication-type="other">Quinteros JA, Markham PF, Lee SW, Hewson KA, Hartley CA, Legione AR, Coppo MJ, Vaz PK, Browning GF. Analysis of the complete genomic sequences of two virus subpopulations of the Australian infectious bronchitis virus vaccine VicS. Avian Pathology. 44(3):182–91.</mixed-citation>
</ref>
<ref id="CR41">
<label>41.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>van Beurden</surname>
<given-names>SJ</given-names>
</name>
<name>
<surname>Berends</surname>
<given-names>AJ</given-names>
</name>
<name>
<surname>Kramer-Kuhl</surname>
<given-names>A</given-names>
</name>
<name>
<surname>Spekreijse</surname>
<given-names>D</given-names>
</name>
<name>
<surname>Chenard</surname>
<given-names>G</given-names>
</name>
<name>
<surname>Philipp</surname>
<given-names>HC</given-names>
</name>
<name>
<surname>Mundt</surname>
<given-names>E</given-names>
</name>
<name>
<surname>Rottier</surname>
<given-names>PJM</given-names>
</name>
<name>
<surname>Verheije</surname>
<given-names>MH</given-names>
</name>
</person-group>
<article-title>Recombinant live attenuated avian coronavirus vaccines with deletions in the accessory genes 3ab and/or 5ab protect against infectious bronchitis in chickens</article-title>
<source>Vaccine</source>
<year>2018</year>
<volume>36</volume>
<issue>8</issue>
<fpage>1085</fpage>
<lpage>1092</lpage>
<pub-id pub-id-type="pmid">29366709</pub-id>
</element-citation>
</ref>
<ref id="CR42">
<label>42.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Benyeda</surname>
<given-names>Z</given-names>
</name>
<name>
<surname>Szeredi</surname>
<given-names>L</given-names>
</name>
<name>
<surname>Mató</surname>
<given-names>T</given-names>
</name>
<name>
<surname>Süveges</surname>
<given-names>T</given-names>
</name>
<name>
<surname>Balka</surname>
<given-names>G</given-names>
</name>
<name>
<surname>Abonyi-Tóth</surname>
<given-names>Z</given-names>
</name>
<name>
<surname>Rusvai</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Palya</surname>
<given-names>V</given-names>
</name>
</person-group>
<article-title>Comparative histopathology and immunohistochemistry of QX-like, Massachusetts and 793/B serotypes of infectious bronchitis virus infection in chickens</article-title>
<source>J Comp Pathol</source>
<year>2010</year>
<volume>143</volume>
<issue>4</issue>
<fpage>276</fpage>
<lpage>283</lpage>
<pub-id pub-id-type="pmid">20570279</pub-id>
</element-citation>
</ref>
<ref id="CR43">
<label>43.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>W-q</surname>
<given-names>F</given-names>
</name>
<name>
<surname>H-n</surname>
<given-names>W</given-names>
</name>
<name>
<surname>Zhang</surname>
<given-names>Y</given-names>
</name>
<name>
<surname>Z-b</surname>
<given-names>G</given-names>
</name>
<name>
<surname>Wang</surname>
<given-names>T</given-names>
</name>
<name>
<surname>C-w</surname>
<given-names>X</given-names>
</name>
<name>
<surname>A-y</surname>
<given-names>Z</given-names>
</name>
<name>
<surname>Yang</surname>
<given-names>X</given-names>
</name>
</person-group>
<article-title>Comparative dynamic distribution of avian infectious bronchitis virus M41, H120, and SAIBK strains by quantitative real-time RT-PCR in SPF chickens</article-title>
<source>Biosci Biotechnol Biochem</source>
<year>2012</year>
<volume>76</volume>
<issue>12</issue>
<fpage>2255</fpage>
<lpage>2260</lpage>
<pub-id pub-id-type="pmid">23221700</pub-id>
</element-citation>
</ref>
<ref id="CR44">
<label>44.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Cheng</surname>
<given-names>J</given-names>
</name>
<name>
<surname>Huo</surname>
<given-names>C</given-names>
</name>
<name>
<surname>Zhao</surname>
<given-names>J</given-names>
</name>
<name>
<surname>Liu</surname>
<given-names>T</given-names>
</name>
<name>
<surname>Li</surname>
<given-names>X</given-names>
</name>
<name>
<surname>Yan</surname>
<given-names>S</given-names>
</name>
<name>
<surname>Wang</surname>
<given-names>Z</given-names>
</name>
<name>
<surname>Hu</surname>
<given-names>Y</given-names>
</name>
<name>
<surname>Zhang</surname>
<given-names>G</given-names>
</name>
</person-group>
<article-title>Pathogenicity differences between QX-like and mass-type infectious bronchitis viruses</article-title>
<source>Vet Microbiol</source>
<year>2018</year>
<volume>213</volume>
<fpage>129</fpage>
<lpage>135</lpage>
<pub-id pub-id-type="pmid">29291996</pub-id>
</element-citation>
</ref>
<ref id="CR45">
<label>45.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Chhabra</surname>
<given-names>R</given-names>
</name>
<name>
<surname>Kuchipudi</surname>
<given-names>SV</given-names>
</name>
<name>
<surname>Chantrey</surname>
<given-names>J</given-names>
</name>
<name>
<surname>Ganapathy</surname>
<given-names>K</given-names>
</name>
</person-group>
<article-title>Pathogenicity and tissue tropism of infectious bronchitis virus is associated with elevated apoptosis and innate immune responses</article-title>
<source>Virology</source>
<year>2016</year>
<volume>488</volume>
<fpage>232</fpage>
<lpage>241</lpage>
<pub-id pub-id-type="pmid">26655241</pub-id>
</element-citation>
</ref>
<ref id="CR46">
<label>46.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Chhabra</surname>
<given-names>R</given-names>
</name>
<name>
<surname>Ball</surname>
<given-names>C</given-names>
</name>
<name>
<surname>Chantrey</surname>
<given-names>J</given-names>
</name>
<name>
<surname>Ganapathy</surname>
<given-names>K</given-names>
</name>
</person-group>
<article-title>Differential innate immune responses induced by classical and variant infectious bronchitis viruses in specific pathogen free chicks</article-title>
<source>Developmental & Comparative Immunology</source>
<year>2018</year>
<volume>87</volume>
<fpage>16</fpage>
<lpage>23</lpage>
<pub-id pub-id-type="pmid">29751011</pub-id>
</element-citation>
</ref>
<ref id="CR47">
<label>47.</label>
<mixed-citation publication-type="other">Bae JY, Lee I, Kim JI, Park S, Yoo K, Park M, Kim G, Park MS, Lee JY, Kang C, et al. A single amino acid in the polymerase acidic protein determines the pathogenicity of influenza B viruses. J Virol. 2018;92(13):e00259–18.</mixed-citation>
</ref>
<ref id="CR48">
<label>48.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Cavanagh</surname>
<given-names>D</given-names>
</name>
<name>
<surname>Davis</surname>
<given-names>PJ</given-names>
</name>
<name>
<surname>Cook</surname>
<given-names>JK</given-names>
</name>
<name>
<surname>Li</surname>
<given-names>D</given-names>
</name>
<name>
<surname>Kant</surname>
<given-names>A</given-names>
</name>
<name>
<surname>Koch</surname>
<given-names>G</given-names>
</name>
</person-group>
<article-title>Location of the amino acid differences in the S1 spike glycoprotein subunit of closely related serotypes of infectious bronchitis virus</article-title>
<source>Avian Pathology</source>
<year>1992</year>
<volume>21</volume>
<issue>1</issue>
<fpage>33</fpage>
<lpage>43</lpage>
<pub-id pub-id-type="pmid">18670913</pub-id>
</element-citation>
</ref>
<ref id="CR49">
<label>49.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Binns</surname>
<given-names>MM</given-names>
</name>
<name>
<surname>Boursnell</surname>
<given-names>ME</given-names>
</name>
<name>
<surname>Tomley</surname>
<given-names>FM</given-names>
</name>
<name>
<surname>Brown</surname>
<given-names>DK</given-names>
</name>
</person-group>
<article-title>Comparison of the spike precursor sequences of coronavirus IBV strains M41 and 6/82 with that of IBV Beaudette</article-title>
<source>J Gen Virol</source>
<year>1986</year>
<volume>67</volume>
<issue>Pt 12</issue>
<fpage>2825</fpage>
<lpage>2831</lpage>
<pub-id pub-id-type="pmid">3025348</pub-id>
</element-citation>
</ref>
<ref id="CR50">
<label>50.</label>
<mixed-citation publication-type="other">Amarasinghe A, Popowich S, De Silva Senapathi U, Abdul-Cader MS, Marshall F, van der Meer F, Cork SC, Gomis S, Abdul-Careem MF. Shell-less egg syndrome (SES) widespread in Western Canadian layer operations is linked to a Massachusetts (mass) type infectious bronchitis virus (IBV) isolate. Viruses. 2018;10(8). 10.3390/v10080437.</mixed-citation>
</ref>
<ref id="CR51">
<label>51.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Stachowiak</surname>
<given-names>BKD</given-names>
</name>
<name>
<surname>Hunton</surname>
<given-names>P</given-names>
</name>
<name>
<surname>Gillingham</surname>
<given-names>S</given-names>
</name>
<name>
<surname>Nagy</surname>
<given-names>É</given-names>
</name>
</person-group>
<article-title>Infectious bronchitis virus surveillance in Ontario commercial layer flocks</article-title>
<source>The Journal of Applied Poultry Research</source>
<year>2005</year>
<volume>14</volume>
<issue>1</issue>
<fpage>141</fpage>
<lpage>146</lpage>
</element-citation>
</ref>
<ref id="CR52">
<label>52.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Ignjatovic</surname>
<given-names>J</given-names>
</name>
<name>
<surname>McWaters</surname>
<given-names>PG</given-names>
</name>
</person-group>
<article-title>Monoclonal antibodies to three structural proteins of avian infectious bronchitis virus: characterization of epitopes and antigenic differentiation of Australian strains</article-title>
<source>J Gen Virol</source>
<year>1991</year>
<volume>72</volume>
<issue>12</issue>
<fpage>2915</fpage>
<lpage>2922</lpage>
<pub-id pub-id-type="pmid">1722501</pub-id>
</element-citation>
</ref>
<ref id="CR53">
<label>53.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Kant</surname>
<given-names>A</given-names>
</name>
<name>
<surname>Koch</surname>
<given-names>G</given-names>
</name>
<name>
<surname>van Roozelaar</surname>
<given-names>DJ</given-names>
</name>
<name>
<surname>Kusters</surname>
<given-names>JG</given-names>
</name>
<name>
<surname>Poelwijk</surname>
<given-names>FAJ</given-names>
</name>
<name>
<surname>van der Zeijst</surname>
<given-names>BAM</given-names>
</name>
</person-group>
<article-title>Location of antigenic sites defined by neutralizing monoclonal antibodies on the S1 avian infectious bronchitis virus glycopolypeptide</article-title>
<source>J Gen Virol</source>
<year>1992</year>
<volume>73</volume>
<issue>3</issue>
<fpage>591</fpage>
<lpage>596</lpage>
<pub-id pub-id-type="pmid">1372036</pub-id>
</element-citation>
</ref>
<ref id="CR54">
<label>54.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Cavanagh</surname>
<given-names>D</given-names>
</name>
<name>
<surname>Davis</surname>
<given-names>PJ</given-names>
</name>
<name>
<surname>Mockett</surname>
<given-names>APA</given-names>
</name>
</person-group>
<article-title>Amino acids within hypervariable region 1 of avian coronavirus IBV (Massachusetts serotype) spike glycoprotein are associated with neutralization epitopes</article-title>
<source>Virus Res</source>
<year>1988</year>
<volume>11</volume>
<issue>2</issue>
<fpage>141</fpage>
<lpage>150</lpage>
<pub-id pub-id-type="pmid">2462314</pub-id>
</element-citation>
</ref>
<ref id="CR55">
<label>55.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Quinteros</surname>
<given-names>JA</given-names>
</name>
<name>
<surname>Lee</surname>
<given-names>S-W</given-names>
</name>
<name>
<surname>Markham</surname>
<given-names>PF</given-names>
</name>
<name>
<surname>Noormohammadi</surname>
<given-names>AH</given-names>
</name>
<name>
<surname>Hartley</surname>
<given-names>CA</given-names>
</name>
<name>
<surname>Legione</surname>
<given-names>AR</given-names>
</name>
<name>
<surname>Coppo</surname>
<given-names>MJC</given-names>
</name>
<name>
<surname>Vaz</surname>
<given-names>PK</given-names>
</name>
<name>
<surname>Browning</surname>
<given-names>GF</given-names>
</name>
</person-group>
<article-title>Full genome analysis of Australian infectious bronchitis viruses suggests frequent recombination events between vaccine strains and multiple phylogenetically distant avian coronaviruses of unknown origin</article-title>
<source>Vet Microbiol</source>
<year>2016</year>
<volume>197</volume>
<fpage>27</fpage>
<lpage>38</lpage>
<pub-id pub-id-type="pmid">27938680</pub-id>
</element-citation>
</ref>
<ref id="CR56">
<label>56.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Abozeid</surname>
<given-names>HH</given-names>
</name>
<name>
<surname>Paldurai</surname>
<given-names>A</given-names>
</name>
<name>
<surname>Khattar</surname>
<given-names>SK</given-names>
</name>
<name>
<surname>Afifi</surname>
<given-names>MA</given-names>
</name>
<name>
<surname>El-Kady</surname>
<given-names>MF</given-names>
</name>
<name>
<surname>El-Deeb</surname>
<given-names>AH</given-names>
</name>
<name>
<surname>Samal</surname>
<given-names>SK</given-names>
</name>
</person-group>
<article-title>Complete genome sequences of two avian infectious bronchitis viruses isolated in Egypt: evidence for genetic drift and genetic recombination in the circulating viruses</article-title>
<source>Infect Genet Evol</source>
<year>2017</year>
<volume>53</volume>
<fpage>7</fpage>
<lpage>14</lpage>
<pub-id pub-id-type="pmid">28495648</pub-id>
</element-citation>
</ref>
<ref id="CR57">
<label>57.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Naguib</surname>
<given-names>MM</given-names>
</name>
<name>
<surname>Höper</surname>
<given-names>D</given-names>
</name>
<name>
<surname>Arafa</surname>
<given-names>A-S</given-names>
</name>
<name>
<surname>Setta</surname>
<given-names>AM</given-names>
</name>
<name>
<surname>Abed</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Monne</surname>
<given-names>I</given-names>
</name>
<name>
<surname>Beer</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Harder</surname>
<given-names>TC</given-names>
</name>
</person-group>
<article-title>Full genome sequence analysis of a newly emerged QX-like infectious bronchitis virus from Sudan reveals distinct spots of recombination</article-title>
<source>Infect Genet Evol</source>
<year>2016</year>
<volume>46</volume>
<fpage>42</fpage>
<lpage>49</lpage>
<pub-id pub-id-type="pmid">27793730</pub-id>
</element-citation>
</ref>
<ref id="CR58">
<label>58.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Wu</surname>
<given-names>X</given-names>
</name>
<name>
<surname>Yang</surname>
<given-names>X</given-names>
</name>
<name>
<surname>Xu</surname>
<given-names>P</given-names>
</name>
<name>
<surname>Zhou</surname>
<given-names>L</given-names>
</name>
<name>
<surname>Zhang</surname>
<given-names>Z</given-names>
</name>
<name>
<surname>Wang</surname>
<given-names>H</given-names>
</name>
</person-group>
<article-title>Genome sequence and origin analyses of the recombinant novel IBV virulent isolate SAIBK2</article-title>
<source>Virus Genes</source>
<year>2016</year>
<volume>52</volume>
<issue>4</issue>
<fpage>509</fpage>
<lpage>520</lpage>
<pub-id pub-id-type="pmid">27108998</pub-id>
</element-citation>
</ref>
<ref id="CR59">
<label>59.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>X-l</surname>
<given-names>L</given-names>
</name>
<name>
<surname>J-l</surname>
<given-names>S</given-names>
</name>
<name>
<surname>J-x</surname>
<given-names>Z</given-names>
</name>
<name>
<surname>G-z</surname>
<given-names>Z</given-names>
</name>
</person-group>
<article-title>Complete genome sequence analysis of a predominant infectious bronchitis virus (IBV) strain in China</article-title>
<source>Virus Genes</source>
<year>2009</year>
<volume>38</volume>
<issue>1</issue>
<fpage>56</fpage>
<lpage>65</lpage>
<pub-id pub-id-type="pmid">18770015</pub-id>
</element-citation>
</ref>
<ref id="CR60">
<label>60.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Khataby</surname>
<given-names>K</given-names>
</name>
<name>
<surname>Souiri</surname>
<given-names>A</given-names>
</name>
<name>
<surname>Kasmi</surname>
<given-names>Y</given-names>
</name>
<name>
<surname>Loutfi</surname>
<given-names>C</given-names>
</name>
<name>
<surname>Ennaji</surname>
<given-names>MM</given-names>
</name>
</person-group>
<article-title>Current situation, genetic relationship and control measures of infectious bronchitis virus variants circulating in African regions</article-title>
<source>The Journal of Basic & Applied Zoology</source>
<year>2016</year>
<volume>76</volume>
<fpage>20</fpage>
<lpage>30</lpage>
</element-citation>
</ref>
<ref id="CR61">
<label>61.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Li</surname>
<given-names>H</given-names>
</name>
<name>
<surname>Yang</surname>
<given-names>H</given-names>
</name>
</person-group>
<article-title>Sequence analysis of nephropathogenic infectious bronchitis virus strains of the Massachusetts genotype in Beijing</article-title>
<source>Avian Pathology</source>
<year>2001</year>
<volume>30</volume>
<issue>5</issue>
<fpage>535</fpage>
<lpage>541</lpage>
<pub-id pub-id-type="pmid">19184943</pub-id>
</element-citation>
</ref>
<ref id="CR62">
<label>62.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Yan</surname>
<given-names>S</given-names>
</name>
<name>
<surname>Liu</surname>
<given-names>X</given-names>
</name>
<name>
<surname>Zhao</surname>
<given-names>J</given-names>
</name>
<name>
<surname>Xu</surname>
<given-names>G</given-names>
</name>
<name>
<surname>Zhao</surname>
<given-names>Y</given-names>
</name>
<name>
<surname>Zhang</surname>
<given-names>G</given-names>
</name>
</person-group>
<article-title>Analysis of antigenicity and pathogenicity reveals major differences among QX-like infectious bronchitis viruses and other serotypes</article-title>
<source>Vet Microbiol</source>
<year>2017</year>
<volume>203</volume>
<fpage>167</fpage>
<lpage>173</lpage>
<pub-id pub-id-type="pmid">28619139</pub-id>
</element-citation>
</ref>
<ref id="CR63">
<label>63.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Khanh</surname>
<given-names>NP</given-names>
</name>
<name>
<surname>Tan</surname>
<given-names>SW</given-names>
</name>
<name>
<surname>Yeap</surname>
<given-names>SK</given-names>
</name>
<name>
<surname>Lee</surname>
<given-names>HJ</given-names>
</name>
<name>
<surname>Choi</surname>
<given-names>KS</given-names>
</name>
<name>
<surname>Hair-Bejo</surname>
<given-names>M</given-names>
</name>
<name>
<surname>Bich</surname>
<given-names>TN</given-names>
</name>
<name>
<surname>Omar</surname>
<given-names>AR</given-names>
</name>
</person-group>
<article-title>Comparative pathogenicity of Malaysian QX-like and variant infectious bronchitis virus strains in chickens at different age of exposure to the viruses</article-title>
<source>J Comp Pathol</source>
<year>2018</year>
<volume>161</volume>
<fpage>43</fpage>
<lpage>54</lpage>
<pub-id pub-id-type="pmid">30173857</pub-id>
</element-citation>
</ref>
<ref id="CR64">
<label>64.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Fulton</surname>
<given-names>RM</given-names>
</name>
<name>
<surname>Reed</surname>
<given-names>WM</given-names>
</name>
<name>
<surname>Thacker</surname>
<given-names>HL</given-names>
</name>
</person-group>
<article-title>Cellular response of the respiratory tract of chickens to infection with Massachusetts 41 and Australian T infectious bronchitis viruses</article-title>
<source>Avian Dis</source>
<year>1993</year>
<volume>37</volume>
<issue>4</issue>
<fpage>951</fpage>
<lpage>960</lpage>
<pub-id pub-id-type="pmid">8141754</pub-id>
</element-citation>
</ref>
<ref id="CR65">
<label>65.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Amarasinghe</surname>
<given-names>A</given-names>
</name>
<name>
<surname>Abdul-Cader</surname>
<given-names>MS</given-names>
</name>
<name>
<surname>Nazir</surname>
<given-names>S</given-names>
</name>
<name>
<surname>De Silva Senapathi</surname>
<given-names>U</given-names>
</name>
<name>
<surname>van der Meer</surname>
<given-names>F</given-names>
</name>
<name>
<surname>Cork</surname>
<given-names>SC</given-names>
</name>
<name>
<surname>Gomis</surname>
<given-names>S</given-names>
</name>
<name>
<surname>Abdul-Careem</surname>
<given-names>MF</given-names>
</name>
</person-group>
<article-title>Infectious bronchitis corona virus establishes productive infection in avian macrophages interfering with selected antimicrobial functions</article-title>
<source>PLoS One</source>
<year>2017</year>
<volume>12</volume>
<issue>8</issue>
<fpage>e0181801</fpage>
<pub-id pub-id-type="pmid">28763472</pub-id>
</element-citation>
</ref>
<ref id="CR66">
<label>66.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Amarasinghe</surname>
<given-names>A</given-names>
</name>
<name>
<surname>Abdul-Cader</surname>
<given-names>MS</given-names>
</name>
<name>
<surname>Almatrouk</surname>
<given-names>Z</given-names>
</name>
<name>
<surname>van der Meer</surname>
<given-names>F</given-names>
</name>
<name>
<surname>Cork</surname>
<given-names>SC</given-names>
</name>
<name>
<surname>Gomis</surname>
<given-names>S</given-names>
</name>
<name>
<surname>Abdul-Careem</surname>
<given-names>MF</given-names>
</name>
</person-group>
<article-title>Induction of innate host responses characterized by production of interleukin (IL)-1beta and recruitment of macrophages to the respiratory tract of chickens following infection with infectious bronchitis virus (IBV)</article-title>
<source>Vet Microbiol</source>
<year>2018</year>
<volume>215</volume>
<fpage>1</fpage>
<lpage>10</lpage>
<pub-id pub-id-type="pmid">29426399</pub-id>
</element-citation>
</ref>
<ref id="CR67">
<label>67.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>da Silva</surname>
<given-names>SEL</given-names>
</name>
<name>
<surname>Ferreira</surname>
<given-names>HL</given-names>
</name>
<name>
<surname>Garcia</surname>
<given-names>AF</given-names>
</name>
<name>
<surname>Silva</surname>
<given-names>FES</given-names>
</name>
<name>
<surname>Gameiro</surname>
<given-names>R</given-names>
</name>
<name>
<surname>Fabri</surname>
<given-names>CUF</given-names>
</name>
<name>
<surname>Vieira</surname>
<given-names>DS</given-names>
</name>
<name>
<surname>Cardoso</surname>
<given-names>TC</given-names>
</name>
</person-group>
<article-title>Mitochondrial bioenergy alterations in avian HD11 macrophages infected with infectious bronchitis virus</article-title>
<source>Arch Virol</source>
<year>2018</year>
<volume>163</volume>
<issue>4</issue>
<fpage>1043</fpage>
<lpage>1049</lpage>
<pub-id pub-id-type="pmid">29302792</pub-id>
</element-citation>
</ref>
<ref id="CR68">
<label>68.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Han</surname>
<given-names>X</given-names>
</name>
<name>
<surname>Tian</surname>
<given-names>Y</given-names>
</name>
<name>
<surname>Guan</surname>
<given-names>R</given-names>
</name>
<name>
<surname>Gao</surname>
<given-names>W</given-names>
</name>
<name>
<surname>Yang</surname>
<given-names>X</given-names>
</name>
<name>
<surname>Zhou</surname>
<given-names>L</given-names>
</name>
<name>
<surname>Wang</surname>
<given-names>H</given-names>
</name>
</person-group>
<article-title>Infectious bronchitis virus infection induces apoptosis during replication in chicken macrophage HD11 cells</article-title>
<source>Viruses</source>
<year>2017</year>
<volume>9</volume>
<issue>8</issue>
<fpage>198</fpage>
</element-citation>
</ref>
<ref id="CR69">
<label>69.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Kameka</surname>
<given-names>AM</given-names>
</name>
<name>
<surname>Haddadi</surname>
<given-names>S</given-names>
</name>
<name>
<surname>Kim</surname>
<given-names>DS</given-names>
</name>
<name>
<surname>Cork</surname>
<given-names>SC</given-names>
</name>
<name>
<surname>Abdul-Careem</surname>
<given-names>MF</given-names>
</name>
</person-group>
<article-title>Induction of innate immune response following infectious bronchitis corona virus infection in the respiratory tract of chickens</article-title>
<source>Virology</source>
<year>2014</year>
<volume>450–451</volume>
<fpage>114</fpage>
<lpage>121</lpage>
</element-citation>
</ref>
<ref id="CR70">
<label>70.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Meir</surname>
<given-names>R</given-names>
</name>
<name>
<surname>Maharat</surname>
<given-names>O</given-names>
</name>
<name>
<surname>Farnushi</surname>
<given-names>Y</given-names>
</name>
<name>
<surname>Simanov</surname>
<given-names>L</given-names>
</name>
</person-group>
<article-title>Development of a real-time TaqMan® RT-PCR assay for the detection of infectious bronchitis virus in chickens, and comparison of RT-PCR and virus isolation</article-title>
<source>J Virol Methods</source>
<year>2010</year>
<volume>163</volume>
<issue>2</issue>
<fpage>190</fpage>
<lpage>194</lpage>
<pub-id pub-id-type="pmid">19781572</pub-id>
</element-citation>
</ref>
<ref id="CR71">
<label>71.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Breslin</surname>
<given-names>JJ</given-names>
</name>
<name>
<surname>Smith</surname>
<given-names>LG</given-names>
</name>
<name>
<surname>Fuller</surname>
<given-names>FJ</given-names>
</name>
<name>
<surname>Guy</surname>
<given-names>JS</given-names>
</name>
</person-group>
<article-title>Sequence analysis of the Turkey coronavirus nucleocapsid protein gene and 3′ untranslated region identifies the virus as a close relative of infectious bronchitis virus</article-title>
<source>Virus Res</source>
<year>1999</year>
<volume>65</volume>
<issue>2</issue>
<fpage>187</fpage>
<lpage>193</lpage>
<pub-id pub-id-type="pmid">10581391</pub-id>
</element-citation>
</ref>
<ref id="CR72">
<label>72.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Klein</surname>
<given-names>D</given-names>
</name>
</person-group>
<article-title>Quantification using real-time PCR technology: applications and limitations</article-title>
<source>Trends Mol Med</source>
<year>2002</year>
<volume>8</volume>
<issue>6</issue>
<fpage>257</fpage>
<lpage>260</lpage>
<pub-id pub-id-type="pmid">12067606</pub-id>
</element-citation>
</ref>
<ref id="CR73">
<label>73.</label>
<element-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Reed</surname>
<given-names>LJ</given-names>
</name>
<name>
<surname>Muench</surname>
<given-names>H</given-names>
</name>
</person-group>
<article-title>A simple method of estimating fifty per cent endpoint</article-title>
<source>Am J Epidemiol</source>
<year>1938</year>
<volume>27</volume>
<issue>3</issue>
<fpage>493</fpage>
<lpage>497</lpage>
</element-citation>
</ref>
</ref-list>
</back>
</pmc>
</record>

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