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Isolation and Metagenomic Identification of Avian Leukosis Virus Associated with Mortality in Broiler Chicken.

ABSTRACT: Avian leukosis virus (ALV) belongs to the family Retroviridae and causes considerable economic losses to the poultry industry. Following an outbreak associated with high mortality in a broiler flock in northern part of Malaysia, kidney tissues from affected chickens were submitted for virus isolation and identification in chicken embryonated egg and MDCK cells. Evidence of virus growth was indicated by haemorrhage and embryo mortality in egg culture. While viral growth in cell culture was evidenced by the development of cytopathic effects. The isolated virus was purified by sucrose gradient and identified using negative staining transmission electron microscopy. Further confirmation was achieved through next-generation sequencing and nucleotide sequence homology search. Analysis of the viral sequences using the NCBI BLAST tool revealed 99-100% sequence homology with exogenous ALV viral envelope protein. Phylogenetic analysis based on partial envelope sequences showed the Malaysian isolate clustered with Taiwanese and Japanese ALV strains, which were closer to ALV subgroup J, ALV subgroup E, and recombinant A/E isolates. Based on these findings, ALV was concluded to be associated with the present outbreak. It was recommended that further studies should be conducted on the molecular epidemiology and pathogenicity of the identified virus isolate.

SUBMITTER: Bande F

PROVIDER: S-EPMC5002465 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Isolation and Metagenomic Identification of Avian Leukosis Virus Associated with Mortality in Broiler Chicken.

Bande Faruku F Arshad Siti Suri SS Omar Abdul Rahman AR

Advances in virology 20160815

Avian leukosis virus (ALV) belongs to the family Retroviridae and causes considerable economic losses to the poultry industry. Following an outbreak associated with high mortality in a broiler flock in northern part of Malaysia, kidney tissues from affected chickens were submitted for virus isolation and identification in chicken embryonated egg and MDCK cells. Evidence of virus growth was indicated by haemorrhage and embryo mortality in egg culture. While viral growth in cell culture was eviden ...[more]

PMID: 27597865

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Project description:BackgroundEndogenous retroviruses (ERVs) are the remnants of retroviral infections which can elicit prolonged genomic and immunological stress on their host organism. In chickens, endogenous Avian Leukosis Virus subgroup E (ALVE) expression has been associated with reductions in muscle growth rate and egg production, as well as providing the potential for novel recombinant viruses. However, ALVEs can remain in commercial stock due to their incomplete identification and association with desirable traits, such as ALVE21 and slow feathering. The availability of whole genome sequencing (WGS) data facilitates high-throughput identification and characterisation of these retroviral remnants.ResultsWe have developed obsERVer, a new bioinformatic ERV identification pipeline which can identify ALVEs in WGS data without further sequencing. With this pipeline, 20 ALVEs were identified across eight elite layer lines from Hy-Line International, including four novel integrations and characterisation of a fast feathered phenotypic revertant that still contained ALVE21. These bioinformatically detected sites were subsequently validated using new high-throughput KASP assays, which showed that obsERVer was highly precise and exhibited a 0% false discovery rate. A further fifty-seven diverse chicken WGS datasets were analysed for their ALVE content, identifying a total of 322 integration sites, over 80% of which were novel. Like exogenous ALV, ALVEs show site preference for proximity to protein-coding genes, but also exhibit signs of selection against deleterious integrations within genes.ConclusionsobsERVer is a highly precise and broadly applicable pipeline for identifying retroviral integrations in WGS data. ALVE identification in commercial layers has aided development of high-throughput diagnostic assays which will aid ALVE management, with the aim to eventually eradicate ALVEs from high performance lines. Analysis of non-commercial chicken datasets with obsERVer has revealed broad ALVE diversity and facilitates the study of the biological effects of these ERVs in wild and domesticated populations.

Project description:BackgroundAvian leukosis virus Subgroup-J (ALV-J) is a rapidly oncogenic evolving retrovirus infecting a variety of avian species; causing severe economic losses to the local poultry industry.MethodsTo investigate ALV-J, a total of 117 blood samples and 57 tissue specimens of different organs were collected for virological, and pathological identification, serological examinations, molecular characterization, and sequencing analysis. To the best of our knowledge, this is the first detailed report recorded in broiler flocks in Egypt. The present study targets the prevalence of a viral tumor disease circulating in broiler flocks in the El-Sharqia, El-Dakahliya, and Al-Qalyubiyya Egyptian governorates from 2021 to 2023 using different diagnostic techniques besides ALV-J gp85 genetic diversity determination.ResultWe first isolated ALV-J on chicken embryo rough cell culture; showing aggregation, rounding, and degeneration. Concerning egg inoculation, embryonic death, stunting, and curling were observed. Only 79 serum samples were positive for ALV-J (67.52%) based on the ELISA test. Histopathological investigation showed tumors consist of uniform masses, usually well-differentiated myelocytes, lymphoid cells, or both in the liver, spleen, and kidneys. Immunohistochemical examination showed that the myelocytomatosis-positive signals were in the spleen, liver, and kidney. The PCR assay of ALV-J gp85 confirmed 545 base pairs with only 43 positive samples (75.4%). Two positive samples were sequenced and submitted to the Genbank with accession numbers (OR509852-OR509853). Phylogenetic analysis based on the gp85 gene showed that the ALV-J Dakahlia-2 isolate is genetically related to ALV-EGY/YA 2021.3, ALV-EGY/YA 2021.4, ALV-EGY/YA 2021.14, and ALV-EGY/YA 2021.9 with amino acid identity percentage 96%, 97%; 96%, 96%; respectively. Furthermore, ALV-J Sharqia-1 isolate is highly genetically correlated to ALV-EGY/YA 2021.14, and ALV-EGY/YA 2021.9, ALV-J isolate QL1, ALV-J isolate QL4, ALV-J isolate QL3, ALV-EGY/YA 2021.4 with amino acid identity percentage 97%, 97%; 98%, 97%, 97%, 95%; respectively.ConclusionsThis study confirmed that ALV-J infection had still been prevalent in broilers in Egypt, and the genetic characteristics of the isolates are diverse.

Dataset Information

Isolation and Metagenomic Identification of Avian Leukosis Virus Associated with Mortality in Broiler Chicken.

Publications

Isolation and Metagenomic Identification of Avian Leukosis Virus Associated with Mortality in Broiler Chicken.

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