aMPV Subtype Research Articles

Avian Metapneumovirus Subtype B Circulation in Poultry and Wild Birds of Colombia.

The global poultry industry, as a leading producer of animal protein, faces significant challenges related to animal health and production due to high bird density and disease risks. A major concern is the Avian Respiratory Complex (ARC), a multifactorial health issue involving pathogens such as avian metapneumovirus (aMPV), an often-underdiagnosed component of the ARC. Wild birds are seen as reservoirs and spreaders of the virus. This study aimed to detect the presence and subtypes of aMPV in samples from breeders, broilers, laying hens, and wild birds in Colombia. A total of 273 samples, including swabs from the upper respiratory and reproductive tracts, were collected from commercial poultry and wild birds. Using nested RT-PCR targeting the G gene, aMPV subtype B was identified in 23 samples (8.42%). Sequencing revealed high genetic similarity to vaccine strains, classifying all viruses as vaccine-like. In the commercial birds, aMPV-B appeared in 21 samples, regardless of symptoms, often in tests for other ARC agents, indicating diagnostic bias. In the wild birds, two samples tested positive, suggesting potential transmission between wild and domestic birds. These findings highlight the need for broader diagnostics and further research into aMPV's impact on avian health.

Detection of Avian Metapneumovirus Subtypes A and B in Moroccan Broiler Farms

Background: Avian metapneumovirus (aMPV) is a widespread infectious respiratory pathogen affecting turkeys and chickens, with co-predominance of the subtypes A and B. Objectives: There is no official reports in Morocco about the subtypes of aMPV circulating. Hence, using quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) subtypes-specific A and B, we aimed at detecting and identifying the potential subtype(s) circulating. Methods: We conducted a longitudinal study on three broiler flocks that were strictly not vaccinated against aMPV and were located in two different geographical regions. We studied two flocks that expressed typical swollen head syndrome (SHS) and sampled them once. Furthermore, we sampled dead birds of one flock confirmed seropositive from a previous study. A total of 118 swabs pooled in 24 samples were subjected to RNA extraction and amplified using a triplex RT-PCR for specific detection of aMPV subtypes A and B. Additionally, serum samples were taken at slaughtering age to cross-check the molecular results. A total of 84 sera were analyzed with a commercial indirect enzyme-linked immunosorbent assay (ELISA) kit to detect and titer antibodies against the two subtypes. Results: Avian metapneumovirus was detected by qRT-PCR in all flocks. About 87.50% of the samples were positive for aMPV-B, and 16.67% for aMPV-A and aMPV-B simultaneously. All flocks showed seropositivity, confirming the molecular findings. Conclusion: The present investigation is the first molecular study in Morocco to elucidate the circulation of aMPV-A and aMPV-B in broiler farms in Morocco with a dominance of aMPV-B and the possibility of co-presence of both subtypes.

Development of an improved reverse genetics system for avian metapneumovirus (aMPV): A novel vaccine vector protects against aMPV and infectious bursal disease virus

Avian metapneumovirus (aMPV), a paramyxovirus, causes acute respiratory diseases in turkeys and swollen head syndrome in chickens. This study established a reverse genetics system for aMPV subtype B LN16-A strain based on T7 RNA polymerase. Full-length cDNA of the LN16-A strain was constructed by assembling five cDNA fragments between the T7 promoter and hepatitis delta virus ribozyme. Transfection of this plasmid, along with the supporting plasmids encoding the N, P, M2-1, and L proteins of LN16-A into BSR-T7/5 cells, resulted in the recovery of aMPV subtype B. To identify an effective insertion site, the enhanced green fluorescent protein (EGFP) gene was inserted into different sites of the LN16-A genome to generate recombinant LN16-As. The results showed that the expression levels of EGFP at the site between the G and L genes of LN16-A were significantly higher than those at the other two sites (between the leader and N genes or replacing the SH gene). To verify the availability of the site between G and L for foreign gene expression, the VP2 gene of very virulent infectious bursal disease virus (vvIBDV) was inserted into this site, and recombinant LN16-A (rLN16A-vvVP2) was successfully rescued. Single immunization of specific-pathogen-free chickens with rLN16A-vvVP2 induced high levels of neutralizing antibodies and provided 100% protection against the virulent aMPV subtype B and vvIBDV. Establishing a reverse genetics system here provides an important foundation for understanding aMPV pathogenesis and developing novel vector vaccines.

Geographical Expansion of Avian Metapneumovirus Subtype B: First Detection and Molecular Characterization of Avian Metapneumovirus Subtype B in US Poultry.

Avian metapneumovirus (aMPV), classified within the Pneumoviridae family, wreaks havoc on poultry health. It typically causes upper respiratory tract and reproductive tract infections, mainly in turkeys, chickens, and ducks. Four subtypes of AMPV (A, B, C, D) and two unclassified subtypes have been identified, of which subtypes A and B are widely distributed across the world. In January 2024, an outbreak of severe respiratory disease occurred on turkey and chicken farms across different states in the US. Metagenomics sequencing of selected tissue and swab samples confirmed the presence of aMPV subtype B. Subsequently, all samples were screened using an aMPV subtype A and B multiplex real-time RT-PCR kit. Of the 221 farms, 124 (56%) were found to be positive for aMPV-B. All samples were negative for subtype A. Six whole genomes were assembled, five from turkeys and one from chickens; all six assembled genomes showed 99.29 to 99.98% nucleotide identity, indicating a clonal expansion event for aMPV-B within the country. In addition, all six sequences showed 97.74 to 98.58% nucleotide identity with previously reported subtype B sequences, e.g., VCO3/60616, Hungary/657/4, and BR/1890/E1/19. In comparison to these two reference strains, the study sequences showed unique 49-62 amino acid changes across the genome, with maximum changes in glycoprotein (G). One unique AA change from T (Threonine) to I (Isoleucine) at position 153 in G protein was reported only in the chicken aMPV sequence, which differentiated it from turkey sequences. The twelve unique AA changes along with change in polarity of the G protein may indicate that these unique changes played a role in the adaptation of this virus in the US poultry. This is the first documented report of aMPV subtype B in US poultry, highlighting the need for further investigations into its genotypic characterization, pathogenesis, and evolutionary dynamics.

Surveillance of Avian Metapneumovirus in Non-Vaccinated Chickens and Co-Infection with Avian Pathogenic Escherichia coli.

Brazil is the second largest producer of broiler chicken in the world, and the surveillance of avian pathogens is of great importance for the global economy and nutrition. Avian metapneumovirus (aMPV) infection results in high rates of animal carcass losses due to aerosacculitis and these impacts can be worsened through co-infection with pathogenic bacteria, particularly Escherichia coli (APEC). The present study evaluated the seroprevalence of the main aMPV subtypes in unvaccinated broiler chickens from poultry farms in Brazil, as well as the clinical effects of co-infection with APEC. Blood samples, respiratory swabs, femurs, liver, and spleen of post-mortem broiler chickens were collected from 100 poultry production batches, totaling 1000 samples. The selection of the production batch was based on the history of systemic and respiratory clinical signs. The results indicated that 20% of the lots showed serological evidence of the presence of aMPV, with two lots being positive for aMPV-B. A total of 45% of batches demonstrated co-infection between aMPV and APEC. The results point to the need for viral surveillance, targeted vaccination, and vaccination programs, which could reduce clinical problems and consequently reduce the use of antibiotics to treat bacterial co-infections.

A novel live attenuated vaccine candidate protects chickens against subtype B avian metapneumovirus

Avian metapneumovirus (aMPV) is a highly contagious pathogen that causes acute upper respiratory tract diseases in chickens and turkeys, resulting in serious economic losses. Subtype B aMPV has recently become the dominant epidemic strain in China. We developed an attenuated aMPV subtype B strain by serial passaging in Vero cells and evaluated its safety and efficacy as a vaccine candidate. The safety test showed that after the 30th passage, the LN16-A strain was fully attenuated, as clinical signs of infection and histological lesions were absent after inoculation. The LN16-A strain did not revert to a virulent strain after five serial passages in chickens. The genomic sequence of LN16-A differed from that of the parent wild-type LN16 (wtLN16) strain and had nine amino acid mutations. In chickens, a single immunization with LN16-A induced robust humoral and cellular immune responses, including the abundant production of neutralizing antibodies, CD4+ T lymphocytes, and the Th1 (IFN-γ) and Th2 (IL-4 and IL-6) cytokines. We also confirmed that LN16-A provided 100% protection against subtype B aMPV and significantly reduced viral shedding and turbinate inflammation. Our findings suggest that the LN16-A strain is a promising live attenuated vaccine candidate that can prevent infection with subtype B aMPV.

Predominance of low pathogenic avian influenza virus H9N2 in the respiratory co-infections in broilers in Tunisia: a longitudinal field study, 2018–2020

Respiratory diseases are a health and economic concern for poultry production worldwide. Given global economic exchanges and migratory bird flyways, respiratory viruses are likely to emerge continuously in new territories. The primary aim of this study was to investigate the major pathogens involved in respiratory disease in Tunisian broiler poultry and their epidemiology. Between 2018 and 2020, broilers farms in northeastern Tunisia were monitored, and 39 clinically diseased flocks were sampled. Samples were screened for five viral and three bacterial respiratory pathogens using a panel of real-time PCR assays. The reemergence of H9N2 low pathogenic avian influenza virus (LPAIV) in commercial poultry was reported, and the Northern and Western African GI lineage strain was typed. The infectious bronchitis virus (IBV) GI-23 lineage and the avian metapneumovirus (aMPV) subtype B also were detected for the first time in broilers in Tunisia. H9N2 LPAIV was the most detected pathogen in the flocks tested, but rarely alone, as 15 of the 16 H9N2 positive flocks were co-infected. Except for infectious laryngotracheitis virus (ILTV), all of the targeted pathogens were detected, and in 61% of the respiratory disease cases, a combination of pathogens was identified. The major combinations were H9N2 + aMPV (8/39) and H9N2 + IBV (6/39), showing the high contribution of H9N2 LPAIV to the multifactorial respiratory diseases. This field survey provided evidence of the emergence of new respiratory viruses and the complexity of respiratory disease in Tunisia. A comprehensive and continuous surveillance strategy therefore is needed to better control respiratory pathogens in Tunisia.

Trends and Challenges in the Surveillance and Control of Avian Metapneumovirus.

Among the respiratory pathogens of birds, the Avian Metapneumovirus (aMPV) is one of the most relevant, as it is responsible for causing infections of the upper respiratory tract and may induce respiratory syndromes. aMPV is capable of affecting the reproductive system of birds, directly impacting shell quality and decreasing egg production. Consequently, this infection can cause disorders related to animal welfare and zootechnical losses. The first cases of respiratory syndromes caused by aMPV were described in the 1970s, and today six subtypes (A, B, C, D, and two more new subtypes) have been identified and are widespread in all chicken and turkey-producing countries in the world, causing enormous economic losses for the poultry industry. Conventionally, immunological techniques are used to demonstrate aMPV infection in poultry, however, the identification of aMPV through molecular techniques helped in establishing the traceability of the virus. This review compiles data on the main aMPV subtypes present in different countries; aMPV and bacteria co-infection; vaccination against aMPV and viral selective pressure, highlighting the strategies used to prevent and control respiratory disease; and addresses tools for viral diagnosis and virus genome studies aiming at improving and streamlining pathogen detection and corroborating the development of new vaccines that can effectively protect herds, preventing viral escapes.

Vaccine Interaction and Protection against Virulent Avian Metapneumovirus (aMPV) Challenge after Combined Administration of Newcastle Disease and aMPV Live Vaccines to Day-Old Turkeys.

Newcastle disease virus (NDV) and avian metapneumovirus (aMPV) are among the most impactful pathogens affecting the turkey industry. Since turkeys are routinely immunized against both diseases, the hatchery administration of the combined respective live vaccines would offer remarkable practical advantages. However, the compatibility of NDV and aMPV vaccines has not yet been experimentally demonstrated in this species. To address this issue, an aMPV subtype B live vaccine was administered to day-old poults either alone or in combination with one of two different ND vaccines. The birds were then challenged with a virulent aMPV subtype B strain, clinical signs were recorded and aMPV and NDV vaccine replication and humoral immune response were assessed. All results supported the absence of any interference hampering protection against aMPV, with no significant differences in terms of clinical scoring. In addition, the mean aMPV vaccine viral titers and antibody titers measured in the dual vaccinated groups were comparable or even higher than in the group vaccinated solely against aMPV. Lastly, based on the NDV viral and antibody titers, the combined aMPV and NDV vaccination does not seem to interfere with protection against NDV, although further studies involving an actual ND challenge will be necessary to fully demonstrate this hypothesis.

First seroepidemiological and risk factor survey of avian metapneumovirus circulation in Moroccan broiler farms

Avian metapneumovirus (aMPV) is a highly contagious pathogen for broilers. Its epidemiological aspects remain poorly understood in Morocco. Given that, we carried out a pilot seroepidemiological survey from December 2020 to June 2021 to define aMPV seroprevalence in Moroccan broiler flocks in different bioclimatic zones (humid to sub-humid, semi-arid, arid) during winter and spring. According to these zones, 48 flocks not vaccinated against aMPV were selected using a stratified random sampling model. With a simple random sampling strategy, fifteen to thirty sera per flock were collected then analysed using a commercial indirect ELISA kit (CIVTEST AVI TRT?, HIPRA S.A., Amer, Spain) that was able to titre antibodies against aMPV subtypes A and B. Furthermore, questionnaires were shared with veterinarians in charge of flocks to collect data and analyse it with multivariable logistic regression models to identify risk factors associated with virus circulation. From 1142 sera, 912 tested positive with the ELISA used, showing an overall aMPV seroprevalence of 79.86% in the broilers. The arid zone had the highest seroprevalences: 94.16% during winter and 84.82% during spring. Bioclimate and season seemed to contribute to aMPV seropositivity. Likewise, high stocking densities and poor hygiene scores on farms were proven to predispose birds to seropositivity. Hereby, we show the circulation of aMPV in Moroccan broiler farms is influenced by bioclimate, season, density, and hygiene conditions. The present study is the first serological evidence of aMPV in broilers in the Maghreb region.

Longitudinal Survey on aMPV Circulation in French Broiler Flocks following Different Vaccination Strategies

In recent years, the impact of respiratory disease resulting from Avian Metapneumovirus (aMPV) infection has been generally rising in the broiler industry in Europe. In this context, in order to investigate aMPV contribution to the clinical picture and the potential benefits of diversified vaccination strategies compared to nonvaccination policies, a longitudinal monitoring was performed, also evaluating Infectious Bronchitis Virus (IBV) presence. Broiler flocks located in Western France, where aMPV has already proven to be a health and productivity issue, were screened by RT-PCR on rhino-pharyngeal swabs, and the viruses were genetically characterized by sequence analysis. For a more comprehensive picture of aMPV molecular epidemiology and evolution in France, aMPV subtype B strains detected from 1985 to 1998 were sequenced and included in the analysis. The survey confirmed the detection of aMPV subtype B in commercial broiler flocks in France, together with a certain heterogeneity demonstrated by the circulation of more recent and historical French field strains. No IBV field strains were detected. The implementation and evaluation of different management choices and vaccine strategies suggests once again that immunization does not prevent infection but contributes greatly to the containment of the clinical manifestations.

Research Note: Evaluation of the cross protective efficacy of a subtype B aMPV vaccine against virulent subtype A aMPV by different administration routes

Avian metapneumovirus (aMPV) causes respiratory and reproductive diseases in birds, including chickens. In the chicken industry, live vaccines against aMPV subtypes A and B, which are the major aMPV subtypes, are widely used to control disease caused by aMPV. In this study, we evaluated the cross protective efficacy of a live aMPV subtype B vaccine administered via 3 different routes (nasal, spray, and oral) against virulent aMPV subtype A in chickens. At 3 wk after vaccination of 1-wk-old specific-pathogen-free chickens, we measured the serological responses. On the same day, we challenged the birds with aMPV subtype A. Protection was evaluated by viral gene detection and histopathological examination at 3 and 5 days postchallenge. Although there were differences in the serological responses according to administration route, all vaccinated birds showed complete protection at 5 days postchallenge. Regardless of administration route, genome of challenge virus was not detected in vaccinated group, and there were significant differences between vaccinated birds and control group. Overall, our results demonstrated that a subtype B aMPV vaccine can provide cross protection against virulent subtype A aMPV in chickens.

Detection and Genome Sequence Analysis of Avian Metapneumovirus Subtype A Viruses Circulating in Commercial Chicken Flocks in Mexico.

Simple SummaryThe use of next-generation sequencing to identify pathogens in clinical samples is continually improving, providing genomic data that can be used to: (1) identify what viruses are present in particular samples, (2) determine the viral genotype, pathotype, or lineage, (3) predict which viruses are likely to cause severe disease, and (4) help guide what vaccines to use for control efforts. As part of a project to improve NGS for diagnostics, samples were collected from commercial chicken farms in Mexico and transported on FTA cards. One of the commonly identified viruses was avian metapneumovirus, an important upper respiratory avian pathogen consisting of subtypes A-D. Complete genomes of seven subtype-A isolates were identified, which are distinct from other previously reported subtype-A strains. Although subtype-A viruses were previously reported in Mexico, this is the first comprehensive complete genome sequence analysis of Mexican subtype-A viruses. These data provide a much clearer picture of viruses circulating in Mexico and demonstrate the value of next-generation sequencing to identify pathogens where surveillance is not routinely performed. Because subtype-A is not found in the U.S., a modified real-time reverse-transcriptase polymerase chain reaction test was evaluated that can quickly identify the virus in poultry samples.Avian metapneumoviruses (aMPV subtypes A-D) are respiratory and reproductive pathogens of poultry. Since aMPV-A was initially reported in Mexico in 2014, there have been no additional reports of its detection in the country. Using nontargeted next-generation sequencing (NGS) of FTA card-spotted respiratory samples from commercial chickens in Mexico, seven full genome sequences of aMPV-A (lengths of 13,288–13,381 nucleotides) were de novo assembled. Additionally, complete coding sequences of genes N (n = 2), P and M (n = 7 each), F and L (n = 1 each), M2 (n = 6), SH (n = 5) and G (n = 2) were reference-based assembled from another seven samples. The Mexican isolates phylogenetically group with, but in a distinct clade separate from, other aMPV-A strains. The genome and G-gene nt sequences of the Mexican aMPVs are closest to strain UK/8544/06 (97.22–97.47% and 95.07–95.83%, respectively). Various amino acid variations distinguish the Mexican isolates from each other, and other aMPV-A strains, most of which are in the G (n = 38), F (n = 12), and L (n = 19) proteins. Using our sequence data and publicly available aMPV-A data, we revised a previously published rRT-PCR test, which resulted in different cycling and amplification conditions for aMPV-A to make it more compatible with other commonly used rRT-PCR diagnostic cycling conditions. This is the first comprehensive sequence analysis of aMPVs in Mexico and demonstrates the value of nontargeted NGS to identify pathogens where targeted virus surveillance is likely not routinely performed.

Molecular Survey on A, B, C and New Avian Metapneumovirus (aMPV) Subtypes in Wild Birds of Northern-Central Italy.

Simple SummaryAvian metapneumovirus (aMPV) is a common pathogen in poultry and has been detected in wild birds, suggesting the possible role in viral dissemination. A feature of aMPV is its genetic and antigenic variability, which has allowed the identification of various subtypes of the virus with different characteristics in terms of host tropism. Two new subtypes of aMPV were recently identified in gulls and parakeets. We aimed to explore the epidemiology of old and new aMPV subtypes in wild birds. Samples were collected in Italy during the surveillance of avian influenza in wild species and were tested with two multiplex real time RT-PCRs that were able to detect and distinguish the aMPV subtypes (A, B, C, gull, and parakeet subtypes). All of the individuals were negative, except for one mallard that was positive for aMPV subtype C. The M and G genes of this strain were molecularly characterized and revealed similarities with Chinese and European strains, including an Italian sequence that was previously detected in a widgeon. These findings confirm the susceptibility of mallards, which are closely related to domestic species, highlighting the importance of the epidemiological monitoring of aMPV circulation.Recent insights into the genetic and antigenic variability of avian metapneumovirus (aMPV), including the discovery of two new subtypes, have renewed interest in this virus. aMPV causes a well-known respiratory disease in poultry. Domestic species show different susceptibility to aMPV subtypes, whereas sporadic detections in wild birds have revealed links between epidemiology and migration routes. To explore the epidemiology of aMPV in wild species, a molecular survey was conducted on samples that were collected from wild birds during avian influenza surveillance activity in Italy. The samples were screened in pools by multiplex real time RT-PCR assays in order to detect and differentiate subtypes A, B, C, and those that have been newly identified. All the birds were negative, except for a mallard (Anas platyrhynchos) that was positive for aMPV subtype C (sampled in Padua, in the Veneto region, in 2018). The sequencing of partial M and full G genes placed the strain in an intermediate position between European and Chinese clusters. The absence of subtypes A and B supports the negligible role of wild birds, whereas subtype C detection follows previous serological and molecular identifications in Italy. Subtype C circulation in domestic and wild populations emphasizes the importance of molecular test development and adoption to allow the prompt detection of this likely emerging subtype.

First detection of avian metapneumovirus subtype C Eurasian lineage in a Eurasian wigeon (Mareca penelope) wintering in Northeastern Italy: an additional hint on the role of migrating birds in the viral epidemiology

ABSTRACT Avian metapneumovirus (aMPV) economically affects the global poultry industry causing respiratory and reproductive disorders. Considering the paucity of data on aMPV occurrence in European free-ranging avifauna, a molecular survey was conducted on wild birds of 23 species belonging to the orders Anseriformes, Charadriiformes or Passeriformes, captured alive and sampled in Northeast Italy as part of the national avian influenza virus (AIV) surveillance activities. A total of 492 oropharyngeal swabs, collected from 2007–2010, all AIV-negative, were screened from aMPV by subtype-specific qRT-PCR. An aMPV-C strain, named aMPV/C/IT/Wigeon/758/07, was found in a wintering young Eurasian wigeon (Mareca penelope) sampled in November 2007. The matrix, fusion, and attachment glycoprotein genes of the detected strain were subsequently amplified by specific independent RT-PCRs, then sequenced, and compared in a phylogenetic framework with known aMPV homologous sequences retrieved from GenBank. Close genetic relationships were found between the aMPV/C/IT/Wigeon/758/07 strain and subtype C Eurasian lineage strains isolated in the late 1990s in French domestic ducks, suggesting epidemiological links. Eurasian wigeons are medium/long-range migrant dabbling ducks that move along the Black Sea/Mediterranean flyway; our finding might, therefore, be related to migratory bridges between countries. To our knowledge, this is the first molecular evidence of the occurrence of aMPV subtype C in Italy and backdates the aMPV-C circulation to 2007. Moreover, the results suggest the susceptibility of Eurasian wigeons to aMPV. Broader investigations are needed to assess the role of wild ducks and the significance of the wildfowl/poultry interface in aMPV-C epidemiology. RESEARCH HIGHLIGHTS Wild birds live-captured in Italy were tested for aMPV detection and characterization. aMPV-C Eurasian lineage was found for the first time in a wintering Eurasian wigeon. Migratory birds could be involved in the aMPV epidemiology.

Nano Zinc Oxide Green-Synthesized from Plumbago auriculata Lam. Alcoholic Extract.

Zinc oxide nanoparticles (ZnO NPs) were synthesized by using an alcoholic extract of the flowering aerial parts of Plumbago auriculata Lam. Dynamic Light Scattering (DLS) revealed that the average size of synthesized ZnO NPs was 10.58 ± 3.350 nm and the zeta potential was −19.6 mV. Transmission electron microscopy (TEM) revealed that the particle size was in the range from 5.08 to 6.56 nm. X-ray diffraction (XRD) analysis verified the existence of pure hexagonal shaped crystals of ZnO nanoparticles with an average size of 35.34 nm in the sample, which is similar to the particle size analysis acquired by scanning electron microscopy (SEM) (38.29 ± 6.88 nm). HPLC analysis of the phenolic ingredients present in the plant extract showed that gallic acid, chlorogenic acid, and catechin were found as major compounds at concentrations of 1720.26, 1600.42, and 840.20 µg/g, respectively. Furthermore, the inhibitory effects of ZnO NPs and the plant extract against avian metapneumovirus (aMPV) subtype B were also investigated. This assessment revealed that the uncalcinated form of Nano-ZnO mediated by P. auriculata Lam. extract possessed a significant antiviral activity with 50% cytotoxic concentration (CC50) and 50% inhibition concentration (IC50) of 52.48 ± 1.57 and 42.67 ± 4.08 µg/mL, respectively, while the inhibition percentage (IP) was 99% and the selectivity index (SI) was 1.23.

Molecular Epidemiology and Genotyping of Infectious Bronchitis Virus and Avian Metapneumovirus in Backyard and Commercial Chickens in Jimma Zone, Southwestern Ethiopia

Poultry production plays a relevant role in the Ethiopian economy and represents a source of poverty alleviation for several social classes. Infectious diseases can therefore significantly impact the economy and welfare. Despite infectious bronchitis virus (IBV) and avian metapneumovirus (aMPV) being present, the knowledge of their epidemiology and impact is extremely limited. In the present work, a cross-sectional study based on 500 tracheal swabs collected from 50 intensive and backyard unvaccinated flocks of the Jimma Zone was performed to investigate the circulation of these viruses and molecularly characterize them. IBV and aMPV presence was tested by molecular assays, and genotyping was carried out on positive samples. Accordingly, 6% (95% CI 2.06% to 16.22%) and 8% (95% CI 3.15% to 18.84%) of flocks tested IBV and aMPV positive, respectively. Particularly, IBV 793B (GI-13) strains were detected in backyard flocks only, and identical or closely related sequences (p-distance <2%) were detected in distantly spaced flocks, suggesting relevant viral circulation. On the contrary, both backyard and intensive flocks were affected by aMPV subtype B. Potential epidemiological links associated to the importation of parental birds from foreign countries could be established. These results highlight non-negligible circulation of these viruses, warranting further epidemiological studies and the evaluation of control measure implementation.

Development of A recombinant nucleocapsid based indirect ELISA for the detection of antibodies to avian metapneumovirus subtypes, A, B, and C

Nucleocapsid (N) protein is the most highly expressed of all avian metapneumovirus (aMPV) viral proteins and stimulates a substantial immune response in infected animals. Codon optimized recombinant N (rec-N) protein from aMPV subtypes A, B, and C were expressed using the baculoviral expression system in Trichoplusia ni (Tni) insect cells. A mixture of purified rec-N antigens from each subtype was used as a coating antigen and was evaluated in indirect ELISA (iELISA) to assess antibody response in serum samples collected from experimentally infected chickens and turkeys with different aMPV subtypes. Also, archived field serum samples that were collected from different poultry submissions were used. Receiver operating characteristic (ROC) analysis was performed using chicken and turkey serum samples that were confirmed by indirect fluorescent antibody (IFA) test for serostatus (positive n = 270, negative n = 610). The ROC analysis showed sensitivity and specificity of 97 % at a cut-off value of 0.25. The rec-N iELISA was compared with a commercial whole virus-based APV kit. The rec-N iELISA showed comparable results in detecting antibody response in aMPV infected chicken sera but was more sensitive in detecting early antibody response in aMPV infected turkey serum samples. Our results further confirm the presence of aMPV antibodies in Canadian domestic poultry populations. The developed aMPV-rec N iELISA offers a safe and valuable alternative to whole virus-based iELISA for serodiagnosis and seroepidemiological surveillance of the disease in domestic poultry.

What is new on molecular characteristics of Avian metapneumovirus strains circulating in Europe?

In the present study, one hundred and sixteen partial G gene sequences of Avian metapneumovirus (aMPV) subtype B, obtained during routine diagnostics in different European Countries in the last few years (2014-2019), were analysed by sequence and phylogenetic analyses in order to draw an updated picture of the molecular characteristics of circulating strains. Nucleotide sequences were compared with other sequences of European and non-European aMPV-Bs collected prior to that period or retrieved from GenBank. Phylogenetic relationships among the aMPV-B strains, reconstructed using the maximum likelihood method implemented in MEGA X, demonstrated that aMPV-B has evolved in Europe from its first appearance, frequently displaying a clear relation with the geographic area of detection. The 40% of aMPV-B viruses analysed were classified as vaccine-derived strains, being phylogenetically related, and showing high nucleotide identity with live commercial vaccine strains licensed in Europe. The remaining 60% were classified as field strains since they clustered separately and showed a low nucleotide identity with vaccines and vaccine-derived strains. The phylogenetic tree showed that the virus has continued to evolve from its first appearance in the '80s since more recently detected strains belonged to clades phylogenetically distant from the older strains. Unlike vaccine-derived strains, field strains tended to cluster according to their geographic origin and irrespective of the host species where the viruses had been detected. In conclusion, the molecular characterization of aMPV-B and the differentiation between vaccines and field strains through G gene sequence analysis can be a useful tool towards correct diagnosis and should be routinely applied in order to better address the control strategies.

Complete Genome Sequence of an Avian Metapneumovirus Subtype B Strain from Hungary.

Avian metapneumoviruses (aMPVs), which have been reported in many countries, cause acute upper respiratory tract disease in chickens and turkeys. Using next-generation sequencing, we report here the complete genome sequence of an aMPV subtype B strain that was isolated from a turkey in Hungary in 1989.