Updated unified phylogenetic classification system and revised nomenclature for Newcastle disease virus

Several Avian paramyxoviruses 1 (synonymous with Newcastle disease virus or NDV, used hereafter) classification systems have been proposed for strain identification and differentiation. These systems pioneered classification efforts; however, they were based on different approaches and lacked objective criteria for the differentiation of isolates. These differences have created discrepancies among systems, rendering discussions and comparisons across studies difficult. Although a system that used objective classification criteria was proposed by Diel and co-workers in 2012, the ample worldwide circulation and constant evolution of NDV, and utilization of only some of the criteria, led to identical naming and/or incorrect assigning of new sub/genotypes. To address these issues, an international consortium of experts was convened to undertake in-depth analyses of NDV genetic diversity. This consortium generated curated, up-to-date, complete fusion gene class I and class II datasets of all known NDV for public use, performed comprehensive phylogenetic neighbor-Joining, maximum-likelihood, Bayesian and nucleotide distance analyses, and compared these inference methods. An updated NDV classification and nomenclature system that incorporates phylogenetic topology, genetic distances, branch support, and epidemiological independence was developed. This new consensus system maintains two NDV classes and existing genotypes, identifies three new class II genotypes, and reduces the number of sub-genotypes. In order to track the ancestry of viruses, a dichotomous naming system for designating sub-genotypes was introduced. In addition, a pilot dataset and sub-trees rooting guidelines for rapid preliminary genotype identification of new isolates are provided. Guidelines for sequence dataset curation and phylogenetic inference, and a detailed comparison between the updated and previous systems are included. To increase the speed of phylogenetic inference and ensure consistency between laboratories, detailed guidelines for the use of a supercomputer are also provided. The proposed unified classification system will facilitate future studies of NDV evolution and epidemiology, and comparison of results obtained across the world

Avian chlamydiosis

B

Highly Pathogenic Avian Influenza (HPAI H5Nx, Clade 2.3.4.4.b) in Poultry and Wild Birds in Sweden: Synopsis of the 2020-2021 Season

Simple Summary Highly pathogenic avian influenza is a virus-induced contagious disease that has killed a large number of poultry and wild birds in Europe in the recent decade and is an increasing problem worldwide. In the winter of 2020-2021, Sweden experienced its worst period to date when the disease was diagnosed on 15 commercial poultry farms and over 2.2 million birds died or were euthanised. The disease was also diagnosed in 130 wild birds and nine flocks of hobby, game or zoo birds between 1 October 2020 and 30 September 2021. The aim of this article was to describe the influenza situation in Sweden during this period and to add to the knowledge related to the alarming situation with highly pathogenic influenza in birds. The disease caused animal suffering and death in wild and domestic birds and incurred high costs due to losses and extensive measures to stop spread. The outbreak investigations, where contacts were traced and virus strains were compared, concluded that the virus was brought to poultry farms by wild birds in most cases. More research is needed to obtain knowledge on risk factors, biosecurity, and wild bird presence on poultry farms to prevent future disease outbreaks. Highly pathogenic avian influenza (HPAI, Gs/Gd lineage) was introduced to Europe in 2005 and has since caused numerous outbreaks in birds. The 2020-2021 season was the hitherto most devastating when considering bird numbers and duration in Europe. Surveillance data, virologic results and epidemiologic investigations from the 2020-2021 outbreaks in Sweden were analysed. Subtypes H5N8 and H5N5 were detected on 24 farms with poultry or other captive birds. In wild birds, subtypes H5N8, H5N5, H5N1, H5N4, H5Nx were detected in 130 out of 811 sampled birds. There was a spatiotemporal association between cases in wild birds and poultry. Based on phylogeny and epidemiology, most of the introductions of HPAI to commercial poultry were likely a result of indirect contact with wild birds. A definite route of introduction to poultry could not be established although some biosecurity breaches were observed. No spread between farms was identified but airborne spread between flocks on the same farm was suspected. Our findings exemplify the challenges posed by the continuously changing influenza viruses that seem to adapt to a broader species spectrum. This points to the importance of wild bird surveillance, compliance to biosecurity, and identification of risk factors for introduction on poultry farms

Investigating the role of poultry in livelihoods and the impact of avian flu on livelihoods outcomes in Africa

In this paper we investigate the role of poultry in households’ livelihoods portfolios and the impact of supply-and-demand shocks that may be caused by highly pathogenic avian influenza (HPAI) on households’ various livelihoods outcomes in four Sub-Saharan African (SSA) countries. The study countries include Ethiopia and Kenya in East Africa and Ghana and Nigeria in West Africa. These countries represent a spectrum of SSA countries regarding disease status, means of disease spread, and the role of the poultry sector in the economy. By using nationally representative household-level secondary data and discrete choice methods (probit and zero-inflated negative binomial models), we profile the household, farm, and regional characteristics of those households that are most likely to keep poultry and those households that are most likely to be engaged in intensive poultry production (that is, to keep larger household flocks). We estimate the ex ante impact of HPAI outbreaks and scares/threats on livelihoods outcomes by using the propensity score matching approach. The results of this study generate valuable information regarding the role of poultry in the livelihoods of small-scale poultry-producing households and the livelihoods impacts of HPAI-induced supply-and-demand shocks. Such information is critical for the design of targeted, and hence effective, HPAI control and mitigation policies.Agricultural growth and technologies, demand shock, highly pathogenic avian influenza (HPAI), Livelihoods, probit model, Propensity score matching, supply shock, zero-inflated negative binomial model,

Prevalencija i molekularna karakterizacija cirkovirusa, astrovirusa i hepadnavirusa u avifauni parka prirode Kopački rit, Hrvatska

Wild birds harbour a plethora of viruses but the research into them has mainly focused on those possessing zoonotic potential or causing significant economic losses in poultry, while others, such as astroviruses (AstVs), circoviruses (CV) and hepadnaviruses (HBV), remain neglected. In this study, 35 dead wild birds belonging to at least 21 species from five orders, collected in Kopački rit Nature Park were tested for the presence of AstVs, CV and HBV using PCR with subsequent nucleotide sequencing. A positive result was found in 18 birds. AstVs were found in three passerine birds, CV was detected in 11 birds belonging to Accipitriformes, Anseriformes, Charadriiformes, Passeriformes or Piciformes orders while HBV was found in four birds belonging to either Anseriformes or Passeriformes. More than one virus was found in a single species (AstV and CV) although in two separate birds. Phylogenetic analysis revealed divergent AstVs somewhat similar to different unclassified AstVs detected in undetermined birds. One AstV shared the highest homology with a bat AstV (98.77%). All CV found in the study resemble avian CV, but none of them clusters in the phylogenetic tree with any avian or avian-like CV sequence. All four HBV sequences detected in this study cluster within the duck HBV group, indicating unusual interspecies transmission to cohabitating passerines. The results of this study represent an important contribution to knowledge about the prevalence, ecology and epidemiology of AstVs, CV and HBV in different wild bird species, indicating the need for further studies, including whole genome sequencing of the detected viruses.Iako divlje ptice nose brojne viruse, istraživanja su uglavnom usmjerena na viruse koji posjeduju zoonotski potencijal ili uzrokuju znatne ekonomske gubitke u peradi, dok ostali, poput astrovirusa (AstV), cirkovirusa (CV) i hepadnavirusa (HBV), ostaju zanemareni. U ovom je istraživanju 35 uginulih divljih ptica, koje pripadaju najmanje 21 vrsti iz pet redova prikupljenih u Parku prirode Kopački rit, testirano na prisutnost AstV-a, CV-a i HBV-a PCR-om uz naknadno nukleotidno sekvenciranje. Pozitivan je rezultat ustanovljen u 18 ptica. AstV je pronađen u tri vrapčarke, CV je otkriven u 11 ptica koje pripadaju redovima Accipitriformes, Anseriformes, Charadriiformes, Passeriformes ili Piciformes dok je HBV pronađen u četiri ptice koje pripadaju rodovima Anseriformes ili Passeriformes. Više od jednog virusa pronađeno je u samo u jednoj vrsti (AstV i CV), iako u dvije zasebne ptice. Filogenetska analiza otkrila je divergentni AstV donekle sličan različitim neklasificiranim astrovirusima otkrivenima u neodređenim vrstama ptica. Jedan AstV dijeli najveću sličnost s AstV-om šišmiša (98,77 %). Svi cirkovirusi pronađeni u istraživanju nalikuju na ptičji CV, ali ni jedan od njih ne grupira se u filogenetskom stablu s bilo kojom sličnom sekvencijom. Sve četiri sekvencije HBV-a otkrivene u ovom istraživanju grupiraju se unutar skupine pačjih hepadnavirusa, što upućuje na neuobičajen prijenos na kohabitirajuće vrapčarke. Rezultati ovog istraživanja važan su doprinos spoznajama o prevalenciji, ekologiji i epidemiologiji AstV-a, CV-a i HBV-a u različitim vrstama divljih ptica, što upućuje na potrebu daljnjih istraživanja, uključujući sekvenciranje cijeloga genoma otkrivenih virusa

Diversidad de patógenos en aves silvestres neotropicales: estrategias de descubrimiento e identificación del papel de las especies hospedadoras

Tesis inédita de la Universidad Complutense de Madrid, Facultad de Ciencias Biológicas, leída el 02/02/2021With this PhD thesis, we want to shed some light on the field of pathogens circulating in wildlife, more specifically, in wild birds. In a global context where pathogen transmission among wildlife, domestic animals and humans is higher than ever before, studies following a discovery-driven approach are essential for human health and biodiversity conservation. However, with some exceptions, few pathogens have been studied in wild birds. In our research, we have focused on two important groups of pathogens carried by this group of animals: viruses and haemosporidians, two models that allow different perspectives in the study of avian pathogens, from discovery to ecological function. Furthermore, we want to highlight the value of remote regions as sources of novel information about pathogen diversity and discovery-driven approaches as essential tools for their study. To this end, we sampled the understory community of wild birds in a tropical rainforest of the Nouragues Natural Reserve (French Guiana) and we analyzed its cloacal virome and the community of haemosporidians infecting them. The general objective of this PhD thesis is to highlight how discovery-driven research on pathogens of wildlife living in remote regions contributes substantially to expand the knowledge in the fields of virology and parasitology. With this information, we will improve the understanding of the diversity, host range, ecology and prevalence of both cloacal viruses and haemosporidians infecting birds from Nouragues. Therefore, the four main objectives of this thesis can be summarized in biodiscovery, screening of pathogens in Guianan birds, highlight remote regions and non-traditional hosts as sources of relevant new information in viral discovery and the analysis of avian malaria parasites in Nouragues...Con esta tesis doctoral se pretende arrojar luz en el campo de los patógenos que circulan en la fauna silvestre, más en concreto, en las aves silvestres. En un contexto global en el que la transmisión de patógenos entre fauna silvestre, animales domésticos y humanos es mayor que nunca, los estudios con aproximaciones enfocadas al descubrimiento de nuevos patógenos en la fauna silvestre son esenciales para la salud humana y la conservación de la biodiversidad. Sin embargo, con algunas excepciones, pocos son los patógenos estudiados en aves silvestres. En nuestra investigación, nos hemos centrado en dos grupos importantes de patógenos que portan este grupo de animales: los virus y los hemosporidios, dos modelos que permiten diferentes perspectivas en el estudio de los patógenos de aves. Además, pretendemos resaltar el valor de las zonas remotas como fuente de nueva información en el ámbito de la diversidad de patógenos y de las aproximaciones enfocadas al descubrimiento como una herramienta esencial para su estudio. Para ello, muestreamos la comunidad de aves silvestres del sotobosque en un bosque tropical húmedo de la Reserva Natural de Nouragues (Guayana Francesa) y analizamos su viroma cloacal y la comunidad de hemosporidios que los infectan. El objetivo general de esta tesis es destacar cómo la investigación dirigida al descubrimiento de nuevos patógenos en fauna silvestre y zonas remotas contribuye sustancialmente a ampliar el conocimiento en los campos de la virología y la parasitología. Con esta información, se mejorará la comprensión de la diversidad, rango de hospedadores, ecología y prevalencia tanto de los virus cloacales como de los hemosporidios que infectan a las aves de Nouragues. Por tanto, los cuatro objetivos principales de esta tesis se resumen en biodescubrimiento, el cribado de patógenos en las aves de Guayana, resaltar los hospedadores no tradicionales y las zonas remotas como fuente de nueva información relevante en el descubrimiento de virus y el análisis de los parásitos de la malaria aviar en Nouragues...Fac. de Ciencias BiológicasTRUEunpu

Tilapia lake virus (TiLV) : utvikling av PCRbaserte diagnostiske metoder og studier av infeksjonsmekanismer

Tilapia lake virus (TiLV) is an emerging virus of wild and farmed tilapiines responsible for causing mortalities and significant economic losses to the aquaculture industry. Since its first report in Israel, the virus has been reported in four continents (Asia, Africa, South and North America) to cause mortalities ranging from the lower extreme of 5-20% and higher extremes of up to 90%. The infection status in many countries is not yet known and implementation of surveillance programs has been recommended. Lake Victoria was selected for TiLV surveillance because it contains a large number of tilapiine species of economic importance to the surrounding East African countries. Well-optimized tools for rapid detection and quantification of the virus are also needed. Moreover, the mode of virus uptake in cells and importance for replication is not known. Therefore, this thesis aimed at understanding the possible presence of TiLV in Lake Victoria in East Africa, to develop tools for detection and quantification of the virus and to shed light of virus uptake mechanisms in permissive cells in vitro. In this thesis, a standard RT-PCR and a quantitative real-time RT-PCR (qRT-PCR) were developed based on all the ten segments of TiLV. The standard PCR was used to screen Nile tilapia (Oreochromis niloticus) from Lake Victoria in Eastern Africa. The quantitative real-time RT-PCR was developed based on virus supernatant of known titre and against samples of unknown virus titre originating from infected TFC #10 cells and fish organs. The virus’ ability to hemagglutinate avian and piscine erythrocytes was assessed, and the modulation of ammonium chloride on uptake and replication of TiLV in E-11 cells was studied. The findings reported in study I showed that primers designed from segment two of TiLV for a standard RT-PCR were the best at detecting TiLV in the infected cells and Nile tilapia organs. TiLV genome was detected in 28 Nile tilapias (14. 66%, N = 191) in which 17.78% (N=45) were from wild fish and 13.70% (N=146) from farmed fish (cage farming). The genomes of circulating TiLV in Nile tilapia from Lake Victoria were identical to those detected from Israel (98%), Ecuador (98%), Thailand (96%), Peru (96%) and USA (97%). TiLV was not grown from infected fish and thus its ability to cause disease in Nile tilapia was not studied. Therefore, I am recommending further studies to fulfil Koch’s postulates. The data reported in study II showed that the developed and optimized quantitative real-time RT-PCR detected TiLV in virus supernatants of known titre and in organs of unknown titre from infected Nile tilapia. The developed assay is sensitive and specific to TiLV with all the primers efficiency being within the range of 95-105%, except primers targeting segment ten that gave an efficiency of 93%. The intra- and inter-assay coefficient of variation ranged between 0.00% ~ 2.63% and 0.00% ~ 5.92%, respectively, which is within the recommended range (below 5%) for an assay to be repeatable and reproducible. The detection limit of 2 TCID50/ml was found for primers targeting segments 1, 2, 3, 4 and 9 while lower detection limit of 20 TCID50/ml was found for primers targeting segment 5, 6, 7, 8 and 10. Overall primers targeting segment 3 had the highest detection limit and primers targeting segment 7 had the lowest detection limit. Interestingly, despite the two primer sets (for segment 3 and 7) having different TiLV detection limits they had an equal amplification efficiency of 98%. Therefore, primer optimization for qRT-PCR is important to optimize assay sensitivity. The study reported in paper III was directed at understanding the hemagglutination property of TiLV using avian and piscine erythrocytes, and the infection mechanisms in E-11 cells. TiLV did not hemagglutinate erythrocytes from any of the species tested indicating that the virus lack hemagglutinin. Further, the study has shown that ammonium chloride does not affect the replication of TiLV in E-11 cells indicating that the virus is not using the endocytic pathway during internalization. Taken together, the two observations suggest that, TiLV is not taken up by receptor-mediated endocytosis during internalization into E-11 cells. Thus, further studies are needed to unravel the uptake mechanism(s), which is the important information for controlling the virus by antiviral agents or immunoprophylaxis.Tilapia lake virus (TiLV) er et fremvoksende virus som infiserer ville arter og oppdrettsarter av tilapia og gir dødelighet og betydelige økonomiske tap i oppdrett. Siden den første beskrivelsen av sykdommen fra Israel, har viruset blitt påvist på fire kontinenter (Asia, Afrika, Sør- og Nord-Amerika) og gir dødelighet fra 5-20% opp mot 90%. Forekomst av viruset er ikke kjent i mange av de landene som driver tilapiaoppdrett, og det er nødvendig å etablere bedre overvåknings- og kontrollprogrammer i disse landene. I denne studien ble Lake Victoria valgt for TiLV-screening fordi den inneholder et stort antall tilapia-arter som er av økonomisk betydning for de omkringliggende østafrikanske landene. Det er også behov for optimaliserte metoder for rask deteksjon og kvantifisering av viruset. Hvordan viruset tas opp i cellene og hvordan det replikerer er ikke kjent. I denne avhandlingen ble det gjennomført studier for å forstå forekomst av TiLV i Lake Victoria i Øst-Afrika, det ble etablert verktøy for påvisning og kvantifisering av viruset med molekylærbiologiske metoder, og det ble gjennomført studier for å bedre forstå opptaksmekanismer i celler under infeksjonen. Det ble utviklet en standard RT-PCR og en kvantitativ RT-PCR (qRT-PCR) basert på alle de ti segmentene til viruset. Standard PCR ble brukt til å undersøke Nile tilapia (Oreochromis niloticus) fra Victoriasjøen. Den kvantitative RT-PCR metoden ble testet mot kjent og ukjent virustiter fra henholdsvis infiserte TFC# 10 celler og organer fra infiserte fisk. Hemagglutinering av røde blodlegemer fra hønsefugl og fisk ble også undersøkt, samt effekten av ammoniumklorid på replikasjonen av TiLV i E-11-celler. Resultatene i studie I viste at primere designet fra segment 2 benyttet for påvisning med standard RT-PCR var best egnet til å påvise TiLV i infiserte cellene og organer fra infisert fisk. TiLV fra Victoriasjøen ble påvist i 28 fisk (14. 66%, N = 191) hvor 17,78% (N = 45) var fra villfisk og 13,70% (N = 146) fra oppdrettsfisk. De sekvensene som ble påvist i Nil-tilapia fra Victoriasjøen var tilnærmet identiske med de som ble påvist i Israel (98%), Ecuador (98%), Thailand (96%), Peru (96%) og USA (97%). TiLV ble ikke dyrket eller testet med tanke på virulens/evne til å forårsake sykdom i Nil-tilapia, og derfor anbefaler jeg videre studier for å oppfylle Kochs postulater. I studie II ble det etablert en ny og optimalisert kvantitativ RT-PCR metode for påvisning av TiLV genom i prøver fra infiserte celler med kjent titer (mengde virus) og fra organer fra infisert Nil-tilapia uten kjent titer. Metoden som ble utviklet er sensitiv og spesifikk for TiLV, og primer-effektiviteten var innenfor et akseptabelt område, 95-105%, bortsett fra primere rettet mot segment 10 (93%). Variasjonskoeffisienten for intra- og inter-analyse varierte mellom henholdsvis 0,00% ~ 2,63% og 0,00% ~ 5,92%, som er innenfor det anbefalte området (under 5%) for at en analyse skal anses som repeterbar og reproduserbar. Sensitiviteten til metoden var 2 TCID50/ml, og primere spesifikke for segmentene 1, 2, 3, 4 og 9 gav samme resultat. En nedre deteksjonsgrense på 20 TCID50/ml ble påvist for primere rettet mot segment 5, 6, 7, 8 og 10. Primere spesifikke for mot segment 3 gav høyest sensitivitet og primere segment 7 den laveste. Begge primersettene hadde en effektivitet på 98%. I artikkel III var målsettingen å forstå hemagglutinasjonsegenskapen til TiLV ved bruk av erythrocytter fra hønsefugl, tilapia og laks, samt betydningen av endocytose i tidlig fase av infeksjonen i E-11-celler. TiLV gir ikke hemagglutinering av erytrocytter fra noen av de testede artene, noe som indikerer at viruset mangler hemagglutinin. Studien har også vist at ammoniumklorid ikke påvirker, dvs. hemmer eller forsinker replikasjonen av TiLV i E-11-celler, noe som indikerer at viruset ikke tas opp ved endocytose. Samlet antyder de to observasjonene at TiLV ikke blir tatt opp av reseptormediert endocytose i E-11-celler. De gjennomførte studiene viser at det er behov for å forstå opptaksmekanismen(e) til virus, som er en viktig informasjonen for å kontrollere virusinfeksjonen med anti-virale midler eller vaksiner

Pathogenesis and Control of Inclusion Body Hepatitis in Broiler chickens

Inclusion body hepatitis (IBH) is an economically important fowl adenovirus (FAdV) disease of broiler chickens. In Canada, FAdV-8a, FAdV-8b, FAdV-11, FAdV-7 and FAdV-2 are the prevalent FAdV serotypes. Currently, there is no commercial vaccine available in Canada to prevent IBH in broiler chickens. The objectives of this study were to develop live, inactivated or subunit FAdV vaccines to control IBH and to identify a suitable adjuvant for an inactivated FAdV vaccine. In chapter 2, we analyzed the efficacy and safety of live and inactivated bivalent FAdV vaccines (FAdV-8b-SK+FAdV-11-1047) against IBH. We demonstrated significant immunoprotection of broiler chickens (98 – 100%) (P<0.01) against IBH by vaccinating broiler breeders with FAdV-8b-SK+FAdV-11-1047 with either a bivalent live vaccine (1x104 TCID50) at 16 weeks of age or a bivalent inactivated vaccine (1x106 TCID50) at 16 and 19 weeks of age. Both the live and inactivated bivalent FAdV vaccines induced broad-spectrum protection against all common serotypes of FAdV circulating in the Canada. Both the live and inactivated FAdV vaccines were equally efficacious in protecting broiler chickens against IBH by passive transfer of maternal antibodies (MtAb) from broiler breeders to their broiler progeny. In chapter 3, we demonstrated that FAdV-8b-SK adjuvanted with CpG-ODN induced a long-lasting humoral immunity similar to inactivated FAdV-8b-SK adjuvanted with Emulsigen-D. FAdV-8b-SK adjuvanted with CpG-ODN induced T helper (Th)-1 and Th-2 type immunity. CpG-ODN as an adjuvant enhanced cytotoxic T-cell memory response of FAdV-8b-SK vaccine. Propagation of some serotypes of FAdVs are difficult in cell lines. Hence, we explored the possibility of developing a subunit FAdV vaccine. In chapter 4, we demonstrated significant protection of broiler chickens against IBH by vaccinating their broiler breeder parents using a FAdV-8b-SK subunit vaccine [fiber protein or virus-like particles (VLPs)]. We also demonstrated that the FAdV-8b-SK fiber and VLPs induce strong cytotoxic T-cell responses in the broiler breeders. The results of this study will help in designing FAdV control strategies for the prevention of IBH in Canada