Vaccine or field strains: the jigsaw pattern of infectious bronchitis virus molecular epidemiology in Poland

Infectious bronchitis (IB), caused by infectious bronchitis virus (IBV), account for severe economic losses in the poultry industry. The continuous emergence of a multitude of IBV variants poses many challenges for its diagnosis and control, and live attenuated vaccines, despite their routine use, still plays a significant role in driving IBV evolution, further complicating the epidemiological scenario. Unfortunately, the impact of different vaccination strategies on IB control, epidemiology, and diagnosis has rarely been investigated. This work presents the results of a large-scale diagnostic survey performed in Poland to study IBV molecular epidemiology and how vaccination may affect the viral circulation in the field. To this purpose, 589 samples were collected between May 2017 and January 2019, tested by reverse transcription-PCR for IBV and sequenced. Vaccine and field strains were discriminated based on genetic and anamnestic information. The most commonly detected lineages were 793B (79%) and variant 2 (17.4%), with sporadic detections of QX, Mass, and D274-like strains. Most of the detected strains had a vaccine origin: 46.3% matched one of the applied vaccines, while 36.5% were genetically related to vaccines not implemented in the respective protocol. Besides their practical value for the proper planning of vaccination protocols in Poland, these results suggest that only a fraction (17.2%) of the circulating strains are field ones, imposing a careful assessment of the actual IBV field menaces. Moreover, phenomena like vaccine spreading and persistence seem to occur commonly, stressing the need to further study the epidemiological consequences of the extensive use of live vaccines

Features of Mild-to-Moderate COVID-19 Patients with Dysphonia

Introduction To explore the prevalence of dysphonia in European patients with mild-to-moderate COVID-19 and the clinical features of dysphonic patients. Methods The clinical and epidemiological data of 702 patients with mild-to-moderate COVID-19 were collected from 19 European Hospitals. The following data were extracted: age, sex, ethnicity, tobacco consumption, comorbidities, general and otolaryngological symptoms. Dysphonia and otolaryngological symptoms were self-assessed through a 4-point scale. The prevalence of dysphonia, as part of the COVID-19 symptoms, was assessed. The outcomes were compared between dysphonic and non-dysphonic patients. The association between dysphonia severity and outcomes was studied through Bayesian analysis. Results A total of 188 patients were dysphonic, accounting for 26.8% of cases. Females developed more frequently dysphonia than males (p=0.022). The proportion of smokers was significantly higher in the dysphonic group (p=0.042). The prevalence of the following symptoms was higher in dysphonic patients compared with non-dysphonic patients: cough, chest pain, sticky sputum, arthralgia, diarrhea, headache, fatigue, nausea and vomiting. The severity of dyspnea, dysphagia, ear pain, face pain, throat pain and nasal obstruction was higher in dysphonic group compared with non-dysphonic group. There were significant associations between the severity of dysphonia, dysphagia and cough. Conclusion Dysphonia may be encountered in a quarter of patients with mild-to-moderate COVID-19 and should be considered as a symptom list of the infection. Dysphonic COVID-19 patients are more symptomatic than non-dysphonic individuals. Future studies are needed to investigate the relevance of dysphonia in the COVID-19 clinical presentation

IMPATTO DI DIFFERENTI STRATEGIE VACCINALI SULLE DINAMICHE DI POPOLAZIONE DEL GENOTIPO QX DI BRONCHITE INFETTIVA E SULLA FREQUENZA DEI FOCOLAI

L’estrema variabilità e la rapida evoluzione del virus della bronchite infettiva (IBV) hanno sempre rappresentato la sfida principale per il suo controllo a causa della limitata cross-protezione fra i diversi ceppi. Vari studi sperimentali hanno dimostrato un incremento nello spettro di protezione qualora gli animali siano vaccinati con più genotipi differenti. Ciò non di meno, le condizioni e la tempistica di somministrazione della vaccinazione in campo sono a tal punto differenti che spesso una generalizzazione dei risultati sperimentali è quantomeno questionabile. Nel presente studio è stato utilizzato un approccio di tipo epidemiologico-filodinamico per ricostruire la storia demografica del principale genotipo di campo (i.e. genotipo QX) circolante in Italia. Centonovantacinque campioni raccolti nel periodo 2012-2016 sono stati sequenziati e analizzati. Ciò ha permesso di evidenziare come sia le dinamiche di popolazione di questo virus che la frequenza di focolai clinici siano state particolarmente variabili nel corso del tempo. Di particolare rilievo è risultata la forte associazione fra questi fenomeni e i cambiamenti nelle strategie vaccinali adottate. Ciò ha permesso di dimostrare il rapporto di causa-effetto fra la somministrazione/ sospensione del vaccino per IBV e le ripercussioni in termini di dimensione della popolazione virale e, soprattutto, dell’occorrenza di episodi clinici indotti da IBV. È stato quindi possibile fornire una robusta dimostrazione dell’effetto protettivo indotto della vaccinazione nella pratica routinaria di allevamento. Inoltre, il resoconto qui riportato evidenzia l’importanza di pianificare con rigore non solo le strategie di controllo ma anche il loro monitoraggio e la loro valutazione, evitando di affidarsi a pareri soggettivi ed estemporanei

First Identification and Molecular Characterization of Avian metapneumovirus Subtype B from Chickens in Greece

Avian metapneumovirus (aMPV) is considered a major pathogen for turkeys but its impact on chicken production is still partially neglected, even though it is fully acknowledged as a primary pathogen in chickens as well. The lack of structured diagnostic surveys does not allow a pervasive understanding of aMPV epidemiology. Being that aMPV is almost an everyday challenge for farmers and veterinarians, a more accurate report of its presence should be detailed, posing the basis for a deep and global epidemiologic analysis. With these premises, the present work aims to report the first detection and molecular characterization of aMPV subtype B field strains from unvaccinated chickens in Greece. The Greek strains appear to be phylogenetically related among each other and with other recent Mediterranean strains while being distant from the currently applied vaccines, thus stressing once more the necessity to evaluate aMPV diffusion and evolution

Viral subpopulation variability in different batches of Infectious bronchitis virus (IBV) vaccines based on GI-23 lineage: Implications for the field

The control of infectious bronchitis (IB) is largely based on routine vaccine administration, often using live-attenuated vaccines. However, their capability to replicate and be transmitted among animals and farms implies significant risks. The detection of strains genetically related to vaccines complicates the diagnostic process and understanding of the viral molecular epidemiology. Moreover, reversion to virulence and associated clinical outbreaks can occur although the underlying mechanism are often unknown. In the present study, three vaccine vials, based on IBV GI-23 lineage (also known as Variant2) were deep sequenced through Next Generation Sequencing (NGS) to investigate the presence and features of viral subpopulations. To elucidate the consequences in the field and identify potential markers suitable for a DIVA strategy, the S1 sequences of strains originating from farms in different countries were sequenced and classified based on the knowledge of their vaccination history and similarity with the applied vaccine. Although all considered vaccine batches shared the same consensus sequence, different subpopulations were identified suggesting independent and poorly constrained evolutionary processes. When compared with strains sampled from farms, the vaccine consensus sequences and the respective subpopulations clustered with vaccine strains and no genetic features were consistently shared with field strains. Therefore, if vaccine-induced outbreaks occur, they are more likely to originate from in vivo evolution rather than selection of already present subpopulations. Although some amino acid residues were most commonly detected in field or vaccine strains, no consistent marker could be identified. The occurrence of subpopulations within IBV GI-23-based vaccines and variability featuring different production batches was demonstrated. Being such a phenomenon apparently driven by random genetic drift rather than directional selection, the differentiation between field and vaccine-derived strains appears extremely challenging based on sequence analysis alone. The knowledge of farm management and vaccination history should thus be considered for a proper epidemiological investigation. © 2022 Elsevier B.V

Molecular epidemiology of infectious bronchitis virus and avian metapneumovirus in Greece

Respiratory diseases like infectious bronchitis virus (IBV) and avian metapneumovirus (aMPV) have been held accountable for major losses for poultry production. Nevertheless, scarce information was present dealing with the prevalence and molecular epidemiology of these infections in Greece and the efficacy of currently applied control strategies. To fill this gap, a specific epidemiological study was designed. A total of 106 broiler and layer farms, including 10 backyard and 96 commercial flocks, were sampled between March 2016 and May 2017, and the obtained tracheal swabs were tested for IBV and aMPV using RT-PCR based techniques followed by sequencing. For each farm, data regarding production type, flock features, clinical signs, and vaccination program were also recorded. Different associations between vaccination protocol, production type, animal category, birds density, age, presence of clinical signs, and IBV and/or aMPV infection were tested. Both IBV and aMPV field strain prevalence were proven high, approximately 20 and 30%, respectively, being the GI-19 lineage (14 out of 19; 73.6%) and B subtype (30 out of 30; 100%), the most commonly detected IBV and aMPV genetic types. Infection with IBV field strains was significantly associated with clinical sign presence (odds ratio = 8.55 [95CI = 2.17–42.90]). Remarkably, only the vaccination protocol involving a double vaccination at 1 D of age was proven protective against IBV-induced symptomatology, with the odds of developing disease being 4.14 [95CI = 1.34–14.51] times lower. No association was demonstrated between aMPV infection and clinical outbreaks or between aMPV and IBV detection, suggesting the marginal role of the former pathogen in poultry farming. Globally, the present study provides the first detailed investigation of the epidemiological scenario of 2 viruses traditionally considered of pivotal relevance in poultry farming and demonstrates that remarkable benefits could be obtained with just minor adjustments in vaccination protocols. © 2019 Poultry Science Association Inc

Vaccine or field strains: the jigsaw pattern of infectious bronchitis virus molecular epidemiology in Poland

Infectious bronchitis (IB), caused by infectious bronchitis virus (IBV), account for severe economic losses in the poultry industry. The continuous emergence of a multitude of IBV variants poses many challenges for its diagnosis and control, and live attenuated vaccines, despite their routine use, still plays a significant role in driving IBV evolution, further complicating the epidemiological scenario. Unfortunately, the impact of different vaccination strategies on IB control, epidemiology, and diagnosis has rarely been investigated. This work presents the results of a large-scale diagnostic survey performed in Poland to study IBV molecular epidemiology and how vaccination may affect the viral circulation in the field. To this purpose, 589 samples were collected between May 2017 and January 2019, tested by reverse transcription-PCR for IBV and sequenced. Vaccine and field strains were discriminated based on genetic and anamnestic information. The most commonly detected lineages were 793B (79%) and variant 2 (17.4%), with sporadic detections of QX, Mass, and D274-like strains. Most of the detected strains had a vaccine origin: 46.3% matched one of the applied vaccines, while 36.5% were genetically related to vaccines not implemented in the respective protocol. Besides their practical value for the proper planning of vaccination protocols in Poland, these results suggest that only a fraction (17.2%) of the circulating strains are field ones, imposing a careful assessment of the actual IBV field menaces. Moreover, phenomena like vaccine spreading and persistence seem to occur commonly, stressing the need to further study the epidemiological consequences of the extensive use of live vaccines. © 2019 The Author(s) 2019. Published by Oxford University Press on behalf of Poultry Science Association

Avian Metapneumovirus subtype B around Europe: a phylodynamic reconstruction

Avian Metapneumovirus (aMPV) has been recognized as a respiratory pathogen of turkey and chickens for a long time. Recently, a crescent awareness of aMPV, especially subtype B, clinical and economic impact has risen among European researchers and veterinarians. Nevertheless, the knowledge of its epidemiology and evolution is still limited. In the present study, the broadest available collection of partial G gene sequences obtained from European aMPV-B strains was analyzed using different phylodynamic and biostatistical approaches to reconstruct the viral spreading over time and the role of different hosts on its evolution. After aMPV-B introduction, approximatively in 1985 in France, the infection spread was relatively quick, involving the Western and Mediterranean Europe until the end of the 1990s, and then spreading westwards at the beginning of the new millennium, in parallel with an increase of viral population size. In the following period, a wider mixing among aMPV-B strains detected in eastern and western countries could be observed. Most of the within-country genetic heterogeneity was ascribable to single or few introduction events, followed by local circulation. This, combined with the high evolutionary rate herein demonstrated, led to the establishment of genetically and phenotypically different clusters among countries, which could affect the efficacy of natural or vaccine-induced immunity and should be accounted for when planning control measure implementation. On the contrary, while a significant strain exchange was proven among turkey, guinea fowl and chicken, no evidence of differential selective pressures or specific amino-acid mutations was observed, suggesting that no host adaptation is occurring

Evaluation of 793/B-like and Mass-like vaccine strain kinetics in experimental and field conditions by real-Time RT-PCR quantification

Infectious bronchitis virus (IBV) is a great economic burden both for productive losses and costs of the control strategies. Many different vaccination protocols are applied in the same region and even in consecutive cycles on the same farm in order to find the perfect balance between costs and benefits. In Northern Italy, the usual second vaccination is more and more often moved up to the chick's first d of life. The second strain administration together with the common Mass priming by spray at the hatchery allows saving money and time and reducing animal stress. The present work compared the different vaccine strains (Mass-like or B48, and 1/96) kinetics both in field conditions and in a 21-day-long experimental trial in broilers, monitoring the viral replication by upper respiratory tract swabbing and vaccine specific real time reverse transcription PCR (RT-PCR) quantification. In both field and experimental conditions, titers for all the vaccines showed an increasing trend in the first 2 wk and then a decrease, though still remaining detectable during the whole monitored period. IBV field strain and avian Metapneumovirus (aMPV) presence also was also investigated by RT-PCR and sequencing, and by multiplex real-Time RT-PCR, respectively, revealing a consistency in the pathogen introduction timing at around 30 d, in correspondence with the vaccine titer's main decrease. These findings suggest the need for an accurate knowledge of live vaccine kinetics, whose replication can compete with the other pathogen one, providing additional protection to be added to what is conferred by the adaptive immune response

Infectious bronchitis virus gel vaccination: evaluation of Mass-like (B-48) and 793/B-like (1/96) vaccine kinetics after combined administration at 1 day of age

Infectious bronchitis (IB) control has a strong impact on poultry farming, because of the necessary epidemiological knowledge for planning the best strategy, the optimal strain association, the priming and boosting interventions. Broiler farming is even more problematic given the short and intense productive cycle, which requires an early onset of protection against most of the infectious threats, possibly with limited respiratory post-vaccination reactions that would have a direct impact on the bird health and productivity. For this purpose, gel vaccination has been proposed as a new approach for infectious bronchitis virus (IBV) control and vaccine intake, kinetics and compatibility of combined strains administered by gel have been analyzed in this study. After gel vaccination with single and combined 1/96 and B-48 strains on 4 groups of commercial broilers, a 21-d-long experimental trial has been conducted to monitor the vaccine safety by clinical assessment and vaccine kinetics by strain-specific real-time RT-PCR on choanal cleft swabs. The vaccine strains administered by gel were safe and negligible respiratory signs were detected, even when combined. Vaccine titers were compared among groups and within the same group among a 10-bird pooled sample and 10 swabs from individually sampled birds. 1/96 strain early reached high titers in all animals, while B-48 presence was less constant even though it was detected in almost all birds before the trial end. The individual and pooled sample comparison revealed a partial overestimation of vaccine titers in the pooled samples and the loss of the prevalence data, although the trend portrayed by the pooled swabs closely followed the individual ones. © 2018 Poultry Science Association Inc