First report of Avian metapneumovirus type B in Iraqi broiler flocks with swollen head syndrome

Background and Aim: Swollen head syndrome (SHS) is a complex disease caused by various agents, including bacterial and viral pathogens, as well as environmental factors. Avian metapneumovirus (aMPV) is one of the most important causes of respiratory diseases and SHS in poultry and one of the most widespread viruses worldwide; however, it has not been recorded in Iraq. This study aimed at the molecular identification and subtyping of aMPV in poultry, with the objectives of investigating the prevalence of aMPV in infected broiler flocks with SHS and molecular typing using primers specific to the study of the prevalence of subtypes A, B, and C of aMPV. Materials and Methods: This study was performed on 67 broiler farms that reported typical SHS from September 2018 to August 2019. Swabs were collected from the trachea, infraorbital sinuses, and lung, then uploaded on FTA cards and subjected to an RNA extraction protocol. Results: aMPV was detected in 16 (23.8%) samples. Molecular typing using primers specific to the attachment glycoprotein (G) gene showed that all positive samples belonged to subtype B, as assessed using the real-time polymerase chain reaction technique. Conclusion: aMPV may be the main etiological factor causing SHS in poultry. Moreover, this was the first report of the prevalence of subtype B aMPV strains in broiler farms in Iraq.

Serological Evidence of Avian Metapneumovirus Infection in Layer Chicken Farms of Madhya Pradesh, India

Background: Avian metapneumovirus (aMPV) is an important viral agent in chicken and turkey production industry and is most commonly associated with acute upper respiratory tract infection and swollen head syndrome. The disease is of significant economic concern as it is highly contagious and can lead to production losses in chicken and turkey flocks, especially when associated with secondary bacterial pathogens. Methods: Herein, we have performed a survey on layer farms in selected areas of Madhya Pradesh State, India to ascertain the aMPV status in laying birds, choanal cleft swabs and sera samples were collected for ELISA and RT-PCR assay. A total of 263 sera and 169 choanal cleft swabs from five districts of Madhya Pradesh State (Indore, Bhopal, Sagar, Guna and Bhind) from layer birds of age group 20-72 weeks were screened for aMPV antibodies and for genome. Result: Out of these Indore had maximum sero-postivity (73.33%), followed by Sagar (5.00%) and Bhopal (4.08%), respectively. Other two districts did not show any seropostivity against aMPV. RT-PCR assay using published primers and in vitro transcribed RNA of nucleoprotein (N) gene as positive control was carried out in 169 choanal cleft samples from four districts (Indore, Bhopal, Sagar and Guna). All the samples were found negative for aMPV genome by RT-PCR assay. Overall, 26/263 (9.88%) sera were positive to aMPV antibodies. As the chickens in India are not vaccinated against avian metapneumovirus therefore, this study indicates that layers are exposed to this important poultry pathogen. This study warrants further investigation in wider geographical area and isolation of aMPV to design the control strategies for aMPV.

New Insights into the Host–Pathogen Interaction of Mycoplasma gallisepticum and Avian Metapneumovirus in Tracheal Organ Cultures of Chicken

Respiratory pathogens are a health threat for poultry. Co-infections lead to the exacerbation of clinical symptoms and lesions. Mycoplasma gallisepticum (M. gallispeticum) and Avian Metapneumovirus (AMPV) are two avian respiratory pathogens that co-circulate worldwide. The knowledge about the host–pathogen interaction of M. gallispeticum and AMPV in the chicken respiratory tract is limited. We aimed to investigate how co-infections affect the pathogenesis of the respiratory disease and whether the order of invading pathogens leads to changes in host–pathogen interaction. We used chicken tracheal organ cultures (TOC) to investigate pathogen invasion and replication, lesion development, and selected innate immune responses, such as interferon (IFN) α, inducible nitric oxide synthase (iNOS) and IFNλ mRNA expression levels. We performed mono-inoculations (AMPV or M. gallispeticum) or dual-inoculations in two orders with a 24-h interval between the first and second pathogen. Dual-inoculations compared to mono-inoculations resulted in more severe host reactions. Pre-infection with AMPV followed by M. gallispeticum resulted in prolonged viral replication, more significant innate immune responses, and lesions (p < 0.05). AMPV as the secondary pathogen impaired the bacterial attachment process. Consequently, the M. gallispeticum replication was delayed, the innate immune response was less pronounced, and lesions appeared later. Our results suggest a competing process in co-infections and offer new insights in disease processes.

Avian Metapneumovirus Subgroup C Induces Mitochondrial Antiviral Signaling Protein Degradation through the Ubiquitin-Proteasome Pathway

The mitochondrial antiviral signaling (MAVS) protein, a critical adapter, links the upstream recognition of viral RNA to downstream antiviral signal transduction. However, the interaction mechanism between avian metapneumovirus subgroup C (aMPV/C) infection and MAVS remains unclear. Here, we confirmed that aMPV/C infection induced a reduction in MAVS expression in Vero cells in a dose-dependent manner, and active aMPV/C replication was required for MAVS decrease. We also found that the reduction in MAVS occurred at the post-translational level rather than at the transcriptional level. Different inhibitors were used to examine the effect of proteasome or autophagy on the regulation of MAVS. Treatment with a proteasome inhibitor MG132 effectively blocked MAVS degradation. Moreover, we demonstrated that MAVS mainly underwent K48-linked ubiquitination in the presence of MG132 in aMPV/C-infected cells, with amino acids 363, 462, and 501 of MAVS being pivotal sites in the formation of polyubiquitin chains. Finally, E3 ubiquitin ligases for MAVS degradation were screened and identified and RNF5 targeting MAVS at Lysine 363 and 462 was shown to involve in MAVS degradation in aMPV/C-infected Vero cells. Overall, these results reveal the molecular mechanism underlying aMPV/C infection-induced MAVS degradation by the ubiquitin-proteasome pathway.

Serological and Molecular Characterization of Avian Metapneumovirus in Chickens in Northern Vietnam

Avian Metapneumovirus (aMPV) is a causative agent of respiratory disease complex in turkeys and chickens that has recently been detected in Vietnam. Due to its novelty, this study was conducted to elucidate the distribution of aMPV in several provinces in northern Vietnam. By the application of Enzyme-Linked Immunosorbent Assay (ELISA) and nested Reverse Transcription-Polymerase Chain Reaction (RT-PCR), this study demonstrated the circulation of aMPV in 12 out of 14 cities/provinces with positive rates of 37.6% and 17.2%, respectively. All nested RT-PCR positive samples were aMPV subgroup B. By pairing the detection results with age groups, it was observed that aMPV infections occurred in chickens of all ages. Additionally, by genetic characterization, aMPV strains were demonstrated to not be attenuated vaccine viruses and to belong to at least two genetic clades. Overall, the obtained results provided insights into the prevalence of aMPV and indicated a greater complexity of respiratory diseases in chickens in Vietnam.

Novel Inactivated Subtype B Avian Metapneumovirus Vaccine Induced Humoral and Cellular Immune Responses

Avian metapneumovirus (aMPV), a highly contagious agent, is widespread and causes acute upper respiratory tract disease in chickens and turkeys. However, currently, there is no vaccine licensed in China. Herein, we describe the development of an inactivated aMPV/B vaccine using the aMPV/B strain LN16. Combined with a novel adjuvant containing immune-stimulating complexes (ISCOMs), the novel vaccine could induce high virus-specific and VN antibodies. In addition, it activated B and T lymphocytes and promoted the expression of IL-4 and IFN-γ. Importantly, boosting vaccination with the inactivated aMPV/B vaccine could provide 100% protection against aMPV/B infection with reduced virus shedding and turbinate inflammation. The protection efficacy could last for at least 6 months. This study yielded a novel inactivated aMPV/B vaccine that could serve as the first vaccine candidate in China, thus contributing to the control of aMPV/B and promoting the development of the poultry industry.

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.