Reannotation of the CELO genome characterizes a set of previously unassigned open reading frames and points to novel modes of host interaction in avian adenoviruses

BACKGROUND: The genome of the avian adenovirus Chicken Embryo Lethal Orphan (CELO) has two terminal regions without detectable homology in mammalian adenoviruses that are left without annotation in the initial analysis. Since adenoviruses have been a rich source of new insights into molecular cell biology and practical applications of CELO as gene a delivery vector are being considered, this genome appeared worth revisiting. We conducted a systematic reannotation and in-depth sequence analysis of the CELO genome. RESULTS: We describe a strongly diverged paralogous cluster including ORF-2, ORF-12, ORF-13, and ORF-14 with an ATPase/helicase domain most likely acquired from adeno-associated parvoviruses. None of these ORFs appear to have retained ATPase/helicase function and alternative functions (e.g. modulation of gene expression during the early life-cycle) must be considered in an adenoviral context. Further, we identified a cluster of three putative type-1-transmembrane glycoproteins with IG-like domains (ORF-9, ORF-10, ORF-11) which are good candidates to substitute for the missing immunomodulatory functions of mammalian adenoviruses. ORF-16 (located directly adjacent) displays distant homology to vertebrate mono-ADP-ribosyltransferases. Members of this family are known to be involved in immuno-regulation and similiar functions during CELO life cycle can be considered for this ORF. Finally, we describe a putative triglyceride lipase (merged ORF-18/19) with additional domains, which can be expected to have specific roles during the infection of birds, since they are unique to avian adenoviruses and Marek's disease-like viruses, a group of pathogenic avian herpesviruses. CONCLUSIONS: We could characterize most of the previously unassigned ORFs pointing to functions in host-virus interaction. The results provide new directives for rationally designed experiments

Re-Annotation Is an Essential Step in Systems Biology Modeling of Functional Genomics Data

One motivation of systems biology research is to understand gene functions and interactions from functional genomics data such as that derived from microarrays. Up-to-date structural and functional annotations of genes are an essential foundation of systems biology modeling. We propose that the first essential step in any systems biology modeling of functional genomics data, especially for species with recently sequenced genomes, is gene structural and functional re-annotation. To demonstrate the impact of such re-annotation, we structurally and functionally re-annotated a microarray developed, and previously used, as a tool for disease research. We quantified the impact of this re-annotation on the array based on the total numbers of structural- and functional-annotations, the Gene Annotation Quality (GAQ) score, and canonical pathway coverage. We next quantified the impact of re-annotation on systems biology modeling using a previously published experiment that used this microarray. We show that re-annotation improves the quantity and quality of structural- and functional-annotations, allows a more comprehensive Gene Ontology based modeling, and improves pathway coverage for both the whole array and a differentially expressed mRNA subset. Our results also demonstrate that re-annotation can result in a different knowledge outcome derived from previous published research findings. We propose that, because of this, re-annotation should be considered to be an essential first step for deriving value from functional genomics data

Genetic analysis of the virulence factors and development of a novel diagnostic antigen and antibody test for Fowl Aviadenovirus

Fowl Aviadenovirus (FAdV) belong to the family Adenoviridae and the genus Aviadenovirus. FAdV is a non-enveloped icosahedral viros which has a linear, double- stranded DNA. It has been grouped into five species and 12 serotypes. FAdVs infections often cause Hydropericardium syndrome (HPS), inclusion body hepatitis (IBH), egg drop syndrome (EDS), gizzard erosion (GE), and respiratory tract disease. The disease occurs worldwide leading to great economic losses in the poultry industry. 　To elucidate factors associated with the virulence of Fowl Aviadenovirus, whole genome sequencing using Next Geneneration Sequencing (NGS) of FAdV A strain JM1/1 was carried out. Developing the non-structural protein based Fluorescent antibody virus neutralization (FAVN) and Western Blotting assay (WB) coupled with Immunofluorescence assay (IFA) were studied. The results shown as following 　1) The complete genome of FAdV A strain JM1/1 is 43,809 nucleotide length. It’s revealed 99% nucleotide sequence identical to FAdV A strain CELO (chicken embryo lethal orphan), which is an European apathogenic reference strain. The nucleotide sequence differences were analyzed, interestingly showing multiple sites insertions and deletions. The results will provide information on the evolution and may help elucidate viral pathogenesis on molecular biology especially on genetic roles. 　2) The recombinant DBP was constructed and subjected as the primary antibody in the developed FAVN test. A DNA Binding Protein (DBP), a non-structural protein, which is detectable early and responsible for initiating DNA replication. DBP was found to be a more conserved domain region within the FAdV serotype 1. The developed FAVN test was compared with a conventional VN test by examining the antibody titer in field chicken sera. The results showed the measured neutralizing antibody titers were high correlated with the VN as the correlation coefficient was 0.8. This FAVN test is simple, achieved quickly, easily and could be an alternative test for FAdV infection. 　3) Likewise, the recombinant 52K was constructed and subjected to use as primary antibody in the developed test based on WB and IFA. 52K, non-structural protein, involve in capsid assembly and/or genome packaging. It is especially expressed in late stages of viral life cycle. The C terminal region of 52K was found to be a more conserved domain within all serotypes. WB and IFA analyze revealed that anti-52K antibody can detect FAdVs infection including homologous and heterologous serotypes. Therefore, this antibody possesses a property which could play a role in an alternative method for FAdVs infection diagnosis. 　In conclusion, this research study provided some knowledge, which will be useful in investigation, surveillance, and elimination of FAdV diseases.博士(獣医学)麻布大