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.

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.

The first isolation and detection of Ornithobacterium rhinotracheale from swollen head syndrome-infected broiler flocks in Iraq.

BACKGROUND AND AIM: The swollen head syndrome (SHS) makes up complex diseases that infect the upper respiratory tract in poultry and causes several economic losses. Furthermore, this syndrome is considered one of the multifactorial etiological agents. Therefore, this study isolated and molecularly detected Ornithobacterium rhinotracheale (ORT) in poultry. MATERIALS AND METHODS: This study was conducted at 67 broiler farms that had birds observed to be infected with the SHS from September 2018 until August 2019. Subsequently, swabs were collected from their trachea, infraorbital sinuses, and lungs, after which obtained samples were treated through two methods: (a) The direct method, by uploading samples on FTA cards, and the indirect method using a transport media. Afterward, reverse transcription-polymerase chain reaction (RT-PCR) was used to analyze the directly treated samples; howeverAQ1, the culture method, followed by PCR, was used to analyze the indirectly treated samples. Next, a partial 16S RNA gene was isolated using four positive PCR products, after which the effect of 16 antibiotics was studied on the seven local ORT strains isolated. RESULTS: The quantity of ORT isolated using the direct method was 28 (41.7%) samples, which were all positive for the strain. Identification was by direct molecular identification (RT-PCR) from samples loaded on FTA cards. Alternatively, 7 (10.4%) ORTs were detected from the indirect method, as obtained using the culture method and biochemical tests. Then, PCR was subsequently used to confirm the results. As observed, 784 bp bands were shown for all seven ORT isolates. Furthermore, results revealed a significant difference in the detection of ORT strains between direct and indirect methods, with p-value (<0.05) and standard deviation of the error±0.038 for the direct, then ±0.061 for the indirect method. For further analysis on the strain types, four 784 bp PCR products were taken, then partial 16S ribosomal sequence typing was conducted. All these four strains were found to be recorded in NCBI for the 1st time as a local Iraqi strain, with accession numbers (MN931657, MN931656, MN931655, and MN931654). Notably, results also showed that all isolated strains were multidrug-resistant. CONCLUSION: From the results, ORT is proposed to be implicated as one of the etiological factors that cause SHSs in poultry. Phylogenetic analysis of the current ORT bacterial strains also showed that they are closely related to the Egyptian isolates.

Genomic sequence and pathogenicity of the first avian metapneumovirus subtype B isolated from chicken in China.

Avian metapneumovirus (aMPV), which has been reported in many countries, causes an acute upper respiratory tract disease in chickens and turkeys. Although aMPV was first detected in China in 1999, there has been no further effort to isolate and characterize the aMPV subtype B (aMPV/B) from field outbreaks. In the present study, we used Vero cells to culture a viral strain, LN16, isolated from chickens with swollen head syndrome. The results of RT-PCR, indirect immunofluorescent antibody, and G gene sequence analyses confirmed that strain LN16 corresponds to aMPV/B. We amplified and sequenced the complete genome of strain LN16 and found it to be 13,513 nucleotides in length. Nine viral protein genes of the strain were between 93.2% and 98.4% identical to those of the pathogenic field isolate VCO3/60616. However, insertions and deletions were detected in the intergenic regions. Animal experiments showed that 72.7% of chickens infected with strain LN16 had excess mucus, nasal discharge, and inflammation in the lungs and turbinate. In addition, 27.2% of chickens infected with LN16 shed progeny virions. Viral tissue distribution analysis showed that aMPV could be detected in the turbinate and occasionally in immune organs. This is the first report of the isolation of aMPV/B in China and the first complete genome sequence of aMPV/B from chicken. These findings enrich the epidemiological data on aMPV and may contribute to the development of effective measures to prevent its further spread in China.

In vivo influence of in vitro up-regulated genes in the virulence of an APEC strain associated with swollen head syndrome.

Avian Pathogenic Escherichia coli is responsible for significant economic losses in the poultry industry by causing a range of systemic or localized diseases collectively termed colibacillosis. The virulence mechanisms of these strains that are pathogenic in poultry and possibly pathogenic in humans have not yet been fully elucidated. This work was developed to study if over-expressed genes in a microarray assay could be potentially involved in the pathogenicity of an Avian Pathogenic Escherichia coli strain isolated from a swollen head syndrome case. For this study, five over-expressed genes were selected for the construction of null mutants [flgE (flagellar hook), tyrR (transcriptional regulator), potF (putrescine transporter), yehD (putative adhesin) and bfr (bacterioferritin)]. The constructed mutants were evaluated for their capacity for the adhesion and invasion of in vitro cultured cells, their motility capacity, and their pathogenic potential in one-day-old chickens compared with the wild-type strain (WT). The Δbfr strain showed a decreased adhesion capacity on avian fibroblasts compared with WT, in the presence and absence of alpha-D-mannopyranoside, and the ΔpotF strain showed decreased adhesion only in the absence of alpha-D-mannopyranoside. The ΔtyrR mutant had a reduced ability to invade Hep-2 cells. No mutant showed changes in invading CEC-32 cells. The mutants ΔflgE and ΔtyrR showed a decreased ability to survive in HD-11 cells. The motility of the mutant strains Δbfr, ΔyehD and ΔpotF was increased, while the ΔtyrR mutant showed reduction, and the ΔflgE became non-motile. No mutant strain caused the same mortality of the WT in one-day-old chickens, showing attenuation to different degrees.

Isolation and characterization of avian metapneumovirus from chickens in Korea

Avian metapneumovirus (aMPV) causes upper respiratory tract infections in chickens and turkeys. Although the swollen head syndrome (SHS) associated with aMPV in chickens has been reported in Korea since 1992, this is the study isolating aMPV from chickens in this country. We examined 780 oropharyngeal swab or nasal turbinate samples collected from 130 chicken flocks to investigate the prevalence of aMPV and to isolate aMPV from chickens from 2004-2008. Twelve aMPV subtype A and 13 subtype B strains were detected from clinical samples by the aMPV subtype A and B multiplex real-time reverse transcription polymerase chain reaction (RRT-PCR). Partial sequence analysis of the G glycoprotein gene confirmed that the detected aMPVs belonged to subtypes A and B. Two aMPVs subtype A out of the 25 detected aMPVs were isolated by Vero cell passage. In animal experiments with an aMPV isolate, viral RNA was detected in nasal discharge, although no clinical signs of SHS were observed in chickens. In contrast to chickens, turkeys showed severe nasal discharge and a relatively higher titer of viral excretion than chickens. Here, we reveal the co-circulation of aMPV subtypes A and B, and isolate aMPVs from chicken flocks in Korea.