Infectious Human Adenoviruses Research Articles

First Evidence of Genotypes Ad3a16 and Ad3a18 in North America, Obtained by Genetic Analysis of Infectious Human Adenovirus from Wastewaters of Two Urban Communities in Canada

A 1-year study found seven infectious human adenovirus serotypes, including Ad3, Ad31, Ad46, Ad27, Ad22, Ad51, and clinical clone PB3, in wastewaters of two major cities in Canada. Comparative genomic analysis revealed the existence of the reportedly highly contagious Ad3a16/18 genotypes. This is the first report of Ad3a16/18 genotypes in North America.

Detection and quantitation of infectious human adenoviruses and JC polyomaviruses in water by immunofluorescence assay

Human adenoviruses (HAdV) and JC polyomaviruses (JCPyV) have been proposed as markers of fecal/urine contamination of human origin. An indirect immunofluorescence assay has been developed to quantify infectious human adenoviruses types 2 and 41 and JC polyomaviruses strain Mad-4 in water samples. The immunofluorescence assay was compared with other quantitative techniques used commonly such as plaque assay, tissue culture infectious dose-50 and quantitative PCR (qPCR). The immunofluorescence assays showed to be specific for the detection of infectious viruses, obtaining negative results when UV or heat-inactivated viruses were analyzed. The assays required less time and showed higher sensitivity for the detection of infectious viral particles than other cell culture techniques (1 log 10 more) evaluated. River water samples spiked previously with human adenoviruses and raw sewage samples were also analyzed using the proposed immunofluorescence assay as well as by qPCR. The results show quantitations with 2 log 10 reduction in the numbers of infectious viruses compared with the number of genome copies detected by qPCR. The immunofluorescence assay developed is fast, sensitive, specific, and a standardizable technique for the quantitation and detection of infectious viruses in water samples.

Quantification of Enteric Viruses, Pathogen Indicators, and Salmonella Bacteria in Class B Anaerobically Digested Biosolids by Culture and Molecular Methods

The most common class B biosolids in the United States are generated by mesophilic anaerobic digestion (MAD), and MAD biosolids have been used for land application. However, the pathogen levels in MAD biosolids are still unclear, especially with respect to enteric viruses. In this study, we determined the occurrence and the quantitative levels of enteric viruses and indicators in 12 MAD biosolid samples and of Salmonella enterica in 6 MAD biosolid samples. Three dewatered biosolid samples were also included in this study for purposes of comparison. Human adenoviruses (HAdV) had the highest gene levels and were detected more frequently than other enteric viruses. The gene levels of noroviruses (NV) reported were comparable to those of enteroviruses (EV) and human polyomaviruses (HPyV). The occurrence percentages of HAdV, HAdV species F, EV, NV GI, NV GII, and HPyV in MAD samples were 83, 83, 42, 50, 75, and 58%, respectively. No hepatitis A virus was detected. Infectious HAdV was detected more frequently than infectious EV, and all infectious HAdV were detected when samples were propagated in A549 cells. Based on most-probable-number (MPN) analysis, A549 cells were more susceptible to biosolid-associated viruses than BGM cells. All indicator levels in MAD biosolids were approximately 10(4) MPN or PFU per gram (dry), and the dewatered biosolids had significantly higher indicator levels than the MAD biosolids. Only two MAD samples tested positive for Salmonella enterica, where the concentration was below 1.0 MPN/4 g. This study provides a broad comparison of the prevalence of different enteric viruses in MAD biosolids and reports the first detection of noroviruses in class B biosolids. The observed high quantitative and infectivity levels of adenoviruses in MAD biosolids indicate that adenovirus is a good indicator for the evaluation of sludge treatment efficiency.

The Possibility of Human Adenovirus Detection From the Conjunctiva in Asymptomatic Cases During Nosocomial Infection

To prevent outbreaks of nosocomial adenoviral conjunctivitis, proper management for transmission control must be performed. We collected conjunctival samples from asymptomatic inpatients and an ophthalmologist in an ophthalmology ward and attempted to detect the human adenovirus (HAdV) pathogen for infection control. One inpatient was diagnosed with adenoviral conjunctivitis on the basis of typical, acute, and severe symptoms and virologic testing by using an immunochromatography (IC) kit. To survey nosocomial infection, conjunctival swabs from 17 other inpatients and 1 ophthalmologist without obvious symptoms of adenoviral conjunctivitis were sampled and analyzed for HAdV pathogens with an IC kit, viral isolation, nested polymerase chain reaction (PCR), and real-time PCR. HAdV antigens and DNA were detected from 1 and 8 of the 18 samples collected for nosocomial survey by IC kit and nested PCR method, respectively. Moreover, infectious HAdV was isolated in the cell culture from only 1 antigen-positive sample. All PCR-positive samples had identical nucleotide sequences of the partial hexon gene and were determined to be HAdV type 37 by phylogenetic analysis. No inpatients tested showed any symptoms of typical adenoviral conjunctivitis, but slight conjunctival infection caused by postoperative reaction and/or mild conjunctivitis that did not resemble HAdV infection was observed. No one developed typical adenoviral conjunctivitis over the 2-month follow-up. The clinical course of adenoviral conjunctivitis varies from inapparent infection to severe conjunctivitis. Mild or inapparent HAdV conjunctival infection could be common during conjunctivitis outbreaks and might play a role in the spread of nosocomial adenoviral conjunctivitis.

Detection of Infectious Human Adenoviruses in Tertiary‐Treated and Ultraviolet‐Disinfected Wastewater

A pilot-scale study investigating the use of low-pressure, high-intensity UV radiation for disinfection of urban wastewater was conducted. The inactivation of coliform bacteria, wastewater-indigenous enteric viruses, seeded poliovirus, and seeded F-specific coliphage was studied. During the course of the pilot study, infectious human adenoviruses were isolated from 15 of 16 large-volume samples of UV-disinfected secondary- and tertiary-treated wastewater. Half of the tertiary-treated, UV-disinfected effluent samples from which the adenoviruses were isolated had total coliform concentrations that complied with California's Water Recycling Criteria. To determine the relative UV resistance of the adenovirus isolates, purified viruses were seeded into tertiary-treated waste-water and exposed to low-pressure, high-intensity, collimated UV radiation. A dose of approximately 170 mW-s/cm2 was required to achieve 99.99% inactivation. These findings suggest that UV doses effective at meeting certain wastewater regulations for total coliform bacteria may not provide suitable inactivation of the UV-resistant human adenoviruses.

A Recombinant Human Adenovirus Vaccine against Rabies

Rabies continues to be a serious problem in both developed and developing nations due to the reservoir of rabies virus in wildlife vectors. The control and worldwide eradication of rabies depends on the development of safe, effective, and economical vaccines that might be used in preexposure vaccination programs for humans and animals. To this end an infectious human adenovirus type 5 recombinant virus that contains the rabies glycoprotein gene, and which may serve as the prototype for a new class of vaccines against rabies, was constructed and tested. This recombinant, when administered by either the parenteral or oronasal route, was highly effective in eliciting good levels of rabies-neutralizing antibodies in the sera of dogs and mice. Mice immunized by the recombinant virus were protected from lethal intracerebral challenge with rabies virus.

A simple technique for the rescue of early region I mutations into infectious human adenovirus type 5.

Early region 1 (E1) of the human adenoviruses has many intriguing properties which have prompted numerous mutational studies to help delineate and characterize the domains responsible for these functions. In mutational analyses being done currently, the E1 region is usually cloned into a bacterial plasmid where it is mutated and then the altered E1 sequences are "rescued" back into infectious virus. The most frequently used rescue procedures are somewhat tedious, requiring the purification and fractionation of linear viral DNA or DNA fragments, and often involve the screening of numerous plaque isolates. Several observations we have made recently on the properties of adenovirus DNA in infected cells and on infectious plasmids in transfected cells led us to design a new approach for rescuing E1 mutations into infectious viral genomes. We constructed a plasmid, pJM17, containing the entire Ad5 DNA molecule, with an insert in the E1 region that exceeds the packaging constraints of the adenovirus capsid. Following transfection of pJM17 into 293 cells the plasmid DNA is able to replicate but cannot be packaged into infectious virions. In contrast cotransfection of 293 cells with pJM17 plus an E1-containing plasmid carrying mutated sequences produces recombinant virions at high efficiencies. Neither plasmid needs to be linearized prior to contransfection. The technique eliminates the need to purify and manipulate infectious virion DNA and since no unique restriction sites are needed, both E1A and E1B mutants' as well as foreign gene inserts in the E1 region can be easily rescued into virus.

Incomplete human adenovirus replication in canine tumour cells and serological evidence of infections in tumour-bearing canine pets

Concentrations of newly synthesized virus antigens and virions in cultures of canine normal and tumour cells after inoculation with equal input multiplicities of human adenovirus types 5, 21 and 31 and infectious canine hepatitis virus (ICHV) were measured by solid phase bead-type radioimmunoassay (SBRIA) and fluorescent antibody techniques with human and canine adenovirus-specific reference sera. Higher concentrations of human adenovirus antigens were detected in inoculated tumour cells than in inoculated normal cells, although no infectious human adenoviruses were detected in either of the inoculated canine cell types. In other cultures of these same canine cells, markedly higher concentrations of both ICHV antigens and progeny virions were detected in infected normal canine kidney cells than in infected neuro-fibrosarcoma cells. Human adenovirus type 5 neutralizing antibody was detected in the sera of 2 of 24 dogs bearing untreated malignant tumours and in the serum of 1 of 20 normal dogs. SBRIA tests showed significantly higher concentrations of human adenovirus antibody in the sera of dogs with tumours than in sera from normal dogs. Concentrations of human Ad5 antibody were greater than those of Ad21 and Ad31 antibody in the sera of both normal and tumour-bearing canine pets, each of which had a history of regular vaccination and of long-term close contact with man. Concentrations of ICHV antibody as measured by virus neutralization and SBRIA tests in sera from normal pet dogs were not significantly different from those detected in sera from tumour-bearing dogs. Supported in part by NIH-NCI-NOI-CB-84 248.