Construction and Characterization of a Novel Recombinant Attenuated and Replication-Deficient Candidate Human Adenovirus Type 3 Vaccine: "Adenovirus Vaccine Within an Adenovirus Vector".

Human adenoviruses (HAdVs) are highly contagious and result in large number of acute respiratory disease (ARD) cases with severe morbidity and mortality. Human adenovirus type 3 (HAdV-3) is the most common type that causes ARD outbreaks in Asia, Europe, and the Americas. However, there is currently no vaccine approved for its general use. The hexon protein contains the main neutralizing epitopes, provoking strong and lasting immunogenicity. In this study, a novel recombinant and attenuated adenovirus vaccine candidate against HAdV-3 was constructed based on a commercially-available replication-defective HAdV-5 gene therapy and vaccine vector. The entire HAdV-3 hexon gene was integrated into the E1 region of the vector by homologous recombination using a bacterial system. The resultant recombinants expressing the HAdV-3 hexon protein were rescued in AD293 cells, identified and characterized by RT-PCR, Western blots, indirect immunofluorescence, and electron microscopy. This potential vaccine candidate had a similar replicative efficacy as the wild-type HAdV-3 strain. However, and importantly, the vaccine strain had been rendered replication-defective and was incapable of replication in A549 cells after more than twenty-generation passages in AD293 cells. This represents a significant safety feature. The mice immunized both intranasally and intramuscularly by this vaccine candidate raised significant neutralizing antibodies against HAdV-3. Therefore, this recombinant, attenuated, and safe adenovirus vaccine is a promising HAdV-3 vaccine candidate. The strategy of using a clinically approved and replication-defective HAdV-5 vector provides a novel approach to develop universal adenovirus vaccine candidates against all the other types of adenoviruses causing ARDs and perhaps other adenovirus-associated diseases.

Household Transmission of Human Adenovirus Type 55 in Case of Fatal Acute Respiratory Disease.

We identified a case of fatal acute respiratory disease from household transmission of human adenovirus type 55 (HAdV-55) in Anhui Province, China. Computed tomography showed severe pneumonia. Comparative genomic analysis of HAdV-55 indicated the virus possibly originated in Shanxi Province, China. More attention should be paid to highly contagious HAdV-55.

Comparative Genomic Analysis of Re-emergent Human Adenovirus Type 55 Pathogens Associated With Adult Severe Community-Acquired Pneumonia Reveals Conserved Genomes and Capsid Proteins.

Human adenovirus type 55 (HAdV-B55) is a recently identified acute respiratory disease (ARD) pathogen in HAdV species B with a recombinant genome between renal HAdV-B11 and respiratory HAdV-B14. Since HAdV-B55 first appeared in China school in 2006, no more ARD cases associated with it had been reported until 2011, when there was an outbreak of adult severe community-acquired pneumonia (CAP) in Beijing, China. Reported here is the bioinformatics analysis of the re-emergent HAdV-B55 responsible for this outbreak. Recombination and protein sequence analysis re-confirmed that this isolate (BJ01) was a recombinant virus with the capsid hexon gene from HAdV-B11. The selection pressures for the three capsid proteins, i.e., hexon, penton base, and fiber genes, were all negative, along with very low non-synonymous (dN) and synonymous (dS) substitutions/site (<0.0007). Phylogenetic analyses of the whole genome and the three major capsid genes of HAdV-B55 revealed the close phylogenetic relationship among all HAdV-B55 strains. Comparative genomic analysis of this re-emergent HAdV-B55 strain (BJ01; 2011) with the first HAdV-B55 strain (QS-DLL; 2006) showed the high genome identity (99.87%), including 10 single-nucleotide non-synonymous substitutions, 11 synonymous substitutions, 3 insertions, and one deletion in non-coding regions. The major non-synonymous substitutions (6 of 10) occurred in the protein pVI in its L3 region, which protein has different functions at various stages of an adenovirus infection, and may be associated with the population distribution of HAdV-B55 infection. No non-synonymous substitutions were found in the three major capsid proteins, which proteins are responsible for type-specific neutralizing antibodies. Comparative genomic analysis of the re-emergent HAdV-B55 strains associated with adult severe CAP revealed conserved genome and capsid proteins, providing the foundation for the development of effective vaccines against this pathogen. This study also facilitates the further investigation of HAdV-B55 epidemiology, molecular evolution, patterns of pathogen emergence and re-emergence, and the predication of genome recombination between adenoviruses.

Comparative genomic analysis of two emergent human adenovirus type 14 respiratory pathogen isolates in China reveals similar yet divergent genomes.

Human adenovirus type 14 (HAdV-B14p) was originally identified as an acute respiratory disease (ARD) pathogen in The Netherlands in 1955. For approximately fifty years, few sporadic infections were observed. In 2005, HAdV-B14p1, a genomic variant, re-emerged and was associated with several large ARD outbreaks across the U.S. and, subsequently, in Canada, the U.K., Ireland, and China. This strain was associated with an unusually higher fatality rate than previously reported for both this prototype and other HAdV types in general. In China, HAdV-B14 was first observed in 2010, when two unrelated HAdV-B14-associated ARD cases were reported in Southern China (GZ01) and Northern China (BJ430), followed by three subsequent outbreaks. While comparative genomic analysis, including indel analysis, shows that the three China isolates, with whole genome data available, are similar to the de Wit prototype, all are divergent from the U.S. strain (303600; 2007). Although the genomes of strains GZ01 and BJ430 are nearly identical, as per their genome type characterization and percent identities, they are subtly divergent in their genome mutation patterns. These genomes indicate possibly two lineages of HAdV-B14 and independent introductions into China from abroad, or subsequent divergence from one; CHN2012 likely represents a separate sub-lineage. Observations of these simultaneously reported emergent strains in China add to the understanding of the circulation, epidemiology, and evolution of these HAdV pathogens, as well as provide a foundation for developing effective vaccines and public health strategies, including nationwide surveillance in anticipation of larger outbreaks with potentially higher fatality rates associated with HAdV-B14p1.

Genome Sequence of a Cynomolgus Macaque Adenovirus (CynAdV-1) Isolate from a Primate Colony in the United Kingdom.

The genome sequence of a simian adenovirus from a cynomolgus macaque, denoted CynAdV-1, is presented here. Phylogenetic analysis supports CynAdV-1 in an independent clade, comprising a new simian adenovirus (SAdV) species. These genome data are critical for understanding the evolution and relationships of primate adenoviruses, including zoonosis and emergent human pathogens.

Molecular Identification and Epidemiological Features of Human Adenoviruses Associated with Acute Respiratory Infections in Hospitalized Children in Southern China, 2012-2013.

BACKGROUND: Acute respiratory infections (ARI) are the major worldwide health problem associated with high morbidity and mortality rates. Human adenovirus (HAdV) is one of the most common pathogens associated with viral ARI, and thus calls for specific diagnosis and better understanding of the epidemiology and clinical characteristics. METHODS: Total 4,130 children with ARI requiring hospitalization from 2012 to 2013 were retrospectively studied. Throat swab specimens were collected from each patient. Fluorescence Quantitative PCR was performed to detect adenovirus as well as other common ARI-related pathogens. The seven HAdV hypervariable regions (HVRs) of the hexon gene from fifty-seven HAdVs-positive samples collected in the seasonal peaks were sequenced. Phylogenetic analysis of HVRs was also conducted to confirm the molecular types and genetic variation. In addition, epidemiological features and co-infection with other human respiratory pathogens were investigated and analyzed. RESULTS: Of 4,130 hospitalized pediatric patients tested, the positive rates of respiratory syncytial virus (RSV), Mycoplasma pneumoniae (MP), and HAdV were 13.7%, 13.2%, and 12.0%, respectively. The HAdV positive patients accounted for 7.9%, 17.2%, 17.5% and 10.7% in age groups <1, 1-3, 3-6 and 6-14 years, respectively. Eighty-four HAdV positive children were co-infected with other respiratory pathogens (84/495, 17.0%). The most common co-infection pathogens with HAdV were MP (57.1%) and Human Bocavirus (HBoV) (16.7%). The majority of HAdV infected patients were totally recovered (96.9%, 480/495); However, four (0.8%) patients, who were previously healthy and at the age of 2 years or younger died of pneumonia. Seasonal peaks of HAdV infection occurred in the summer season of 2012 and 2013; the predominant HAdV type was HAdV-3 (70%), followed by HAdV-7 (28%). These epidemiological features were different from those in Northern China. The HAdV-55 was identified and reported for the first time in Guangzhou metropolitan area. Phylogenetic analysis indicated that all the HVR sequences of the hexon gene of HAdV-3 and -7 strains have high similarity within their individual types, and these strains were also similar to those circulating in China currently, indicating the conservation of hexon genes of both HAdV-3 and HAdV-7. CONCLUSIONS: Knowledge of the epidemiological features and molecular types of HAdV, a major pathogen of pediatric ARI, as well as other co-infected respiratory pathogens circulating in Guangzhou, southern China, is vital to predict and prevent future disease outbreaks in children. This study will certainly facilitate HAdV vaccine development and treatment of HAdV infections in children.

Re-emergent human adenovirus genome type 7d caused an acute respiratory disease outbreak in Southern China after a twenty-one year absence.

Human adenoviruses (HAdVs) are highly contagious pathogens causing acute respiratory disease (ARD), among other illnesses. Of the ARD genotypes, HAdV-7 presents with more severe morbidity and higher mortality than the others. We report the isolation and identification of a genome type HAdV-7d (DG01_2011) from a recent outbreak in Southern China. Genome sequencing, phylogenetic analysis, and restriction endonuclease analysis (REA) comparisons with past pathogens indicate HAdV-7d has re-emerged in Southern China after an absence of twenty-one years. Recombination analysis reveals this genome differs from the 1950s-era prototype and vaccine strains by a lateral gene transfer, substituting the coding region for the L1 52/55 kDa DNA packaging protein from HAdV-16. DG01_2011 descends from both a strain circulating in Southwestern China (2010) and a strain from Shaanxi causing a fatality and outbreak (Northwestern China; 2009). Due to the higher morbidity and mortality rates associated with HAdV-7, the surveillance, identification, and characterization of these strains in population-dense China by REA and/or whole genome sequencing are strongly indicated. With these accurate identifications of specific HAdV types and an epidemiological database of regional HAdV pathogens, along with the HAdV genome stability noted across time and space, the development, availability, and deployment of appropriate vaccines are needed.