Relevant Viruses Research Articles

Live virus survives excimer laser ablation

Objectives To determine whether live virus can withstand excimer laser ablation and pose a possible health hazard to medical personnel. Design Experimental study. Methods Fibroblasts infected with oral polio vaccine virus were ablated with an excimer laser. The plume was collected using a smoke evacuator and bubbled through viral culture media. Main outcome measures The inlet tube from the smoke evacuator was swabbed and cultured for virus. Liquid from the bubble trap was also cultured. Results Live virus was shown in the material trapped from the laser plume. Conclusions Oral polio vaccine virus can survive excimer laser ablation. Whether other more clinically relevant viruses, such as human immunodeficiency virus, can withstand ablation and remain infectious remains to be determined.

Inactivation Kinetics of Model and Relevant Blood-borne Viruses by Treatment with Sodium Hydroxide and Heat

To determine the efficacy of a clean-in-place system for the inactivation of viruses present in human plasma, the effect of 0·1Msodium hydroxide at 60°C on viral infectivity was investigated. Inactivation of the following model and relevant viruses were followed as a function of time: human hepatitis A virus (HAV), canine parvovirus (CPV; a model for human parvovirus B-19) pseudorabies virus (PRV, a model for hepatitis B virus), and bovine viral diarrhoea virus (BVDV, a model for hepatitis C virus and human immunodeficiency virus). Infectivity of CPV was determined by a novelin situEIA method which will prove useful for studies to validate parvovirus inactivation or removal. Infectivity of BVDV, PRV and CPV were shown to be reproducibly inactivated below the limit of detection by 0·1MNaOH at 60°C within 30s. HAV was inactivated to below the limit of detection within 2 min. Treatment with heat alone also resulted in some log reduction for all viruses tested except for CPV which remained unaffected after heating at 60°C for 16 min. Treatment of HAV with hydroxide alone (up to 1·0m) at 15°C did not lead to rapid inactivation. Collectively, these data suggest that 0·1MNaOH at 60°C for two min should be sufficient to inactivate viruses present in process residues.

Inactivation of viruses by β-propiolactone in human cyro poor plasma and IgG concentrates

Virus inactivation by cold treatment with β-propiolactone (BPL) was investigated in human cyro poor plasma and purified IgG concentrates spiked with relevant human viruses or appropriate animal model viruses.The samples were treated with 0.1 or 0.25% BPL for 300 or 480 min, respectively. Residual infectivity was determined by standard microtitration assays on tissue culture cells. The inactivation of all viruses tested was more effective in IgG than in plasma. IgG: R1=4–5.5 log10for vesicular stomatitis virus (VSV). Semliki Forest virus (SFV), bovine virus diarrhoea virus (BVDV), murine encephalomyelitis virus (MEV), feline calicivirus (FVC), suid parvovirus (PPV), simian virus 40 (SV40); R1=2–4 log10for suid herpesvirus type 1 (SHV-1), bovine herpesvirus type 1 (BHV-1), human immunodeficiency virus type 2 (HIV-2), simian immunodeficiency virus (SIVagm3). Plasma: R1=3–5 log10for VSV, SFV, BVDV, SHV-1, MEV:R1=0–3 log10for HIV-1, SIVagm3BHV-1, FCV, PPV, SV40. After addition of SIVagm3, HIV-2, and PPV to plasma or lgG, spontaneous inactivation without further addition of BPL was observed.These results demonstrate that treatment with BPL has a limited capacity to inactivate viruses. Different inactivation kinetics were observed in plasma and IgG concentrates. Therefore, virus inactivation by BPL must be tested for individual blood products independently and should not be extrapolated from other model systems.

The prevalence of indices of hepatitis C and B infection, and elevated aminotransferase enzymes in patients with oral lichen planus (OLP) in eastern Saudi Arabia

To determine the possibility of any association between HBV, HCV, elevated aminotransferase enzymes and Oral Lichen Planus (OLP)patients in Eastern Saudi Arabia. Sera were collected from OLP patients, to be tested for HbsAg, anti-HCV and ALT/AST levels. All the patients who were diagnosed clinically in periodontal section, Dammam Central Hospital were from Eastern Saudi Arabia. The histopathological diagnosis was done in Histopathology Section, Dammam Regional Labs, the virological studies in the Virus Diagnosis Lab of Dammam Regional Labs and Blood Bank, and the aminotransferase tests done in the Dammam Central Hospital Labs. 34 serum specimens were collected from OLP patients, and 32 other samples from healthy populations of the same age and sex as the controls of the study. Incidence of HBsAg, anti-HCV, HBsAg+HCV, and elevated aminotransferase enzymes among OLP patients were 8.8%, 14.7%, 2.9%, and 47.05% respectively and the results from the control subjects were 6.25%, 3.12%, 0%, and 3.12% respectively. There is a significant association between OLP and HCV infection. No clear evidence of this relationship appeared with HBV. All the aminotransferase elevated samples were positive to HCV, giving a clear evidence of the association of chronic HCV infection with the OLP. Aminotransferase elevated results could be used as a clue to clinical signs of asymptomatic hepatopathies, and as a marker to check the OLP cases for the relevant Hepatic Viruses. Despite the limited number of OLP patients in this study, the results could highlight the problem in this geographical area of the world. We recommend a comprehensive study to be carried out using this current study as a preliminary one.

Solvent/detergent treatment of human plasma--a very robust method for virus inactivation. Validated virus safety of OCTAPLAS.

OCTAPLAS is a cell-free standardized blood group specific human coagulation active plasma for transfusion. The viral safety is mainly based on the treatment with solvent/detergent. SD is known to irreversibly inactivate the lipid enveloped viruses including HIV 1 + 2, HBV and HCV and has been recommended both by European guidelines and the FDA. The potential limitation of each inactivation method should be validated according to the current EU-guidelines CPMP/BWP/268/95 and CPMP/BWP/269/95. Our studies demonstrate that the SD method inactivates lipid enveloped viruses within a few minutes to below the limit of detection. The concentration of the SD reagents as well as several other process parameters chosen for the virus inactivation step of plasma pools of different protein and lipid composition result in a high safety margin towards the clinically relevant viruses.

Antiserum inhibition of propagating viruses

The design and implementation of controlled environments to continuously culture and evolve viruses provides a means to track how their populations respond to natural and designed anti-viral agents. We have previously demonstrated how the growth of viruses in spreading plaques enables detection and characterization of their evolutionary dynamics. Using plaques of phage T7 growing on E. coli as a model system, we observe here that velocities of propagation can be readily controlled by the level of anti-viral antiserum incorporated into the propagation medium. Further, we develop a simple analytic expression for the radial velocity of propagation in terms of the microscopic rates of viral amplification, Fickian diffusion of the virions and their neutralization by antiserum. Our analysis captures the essential dependence of propagation velocity on antiserum concentration. This study provides an ex vivo foundation for exploring how medically relevant viruses escape suppression by the immune system.

Monovalent single-chain Fv fragments and bivalent miniantibodies bound to vesicular stomatitis virus protect against lethal infection.

Several antibody-dependent mechanisms have been postulated to mediate neutralization of different animal viruses, including blocking of docking to receptors, induction of conformational changes in the virus coat, and Fc-dependent opsonization. We have studied the molecular requirements for antibody-mediated neutralization of vesicular stomatitis virus (VSV) in vitro and protection against lethal disease in vivo with a single-chain Fv fragment (scFv) and the corresponding bivalent miniantibody (scFv-dHLX) generated from a VSV-neutralizing monoclonal antibody. Both monovalent scFv and bivalent scFv-dHLX miniantibodies were able to neutralize VSV in vitro and to protect interferon-alphabeta receptor-deficient (IFN-alphabeta R-/-) mice against lethal disease after intravenous injection of 50 plaque-forming units (pfu) VSV pre-incubated with the scFv reagents. Similarly, severe-combined immunodeficient (SCID) mice infected with immune complexes of 10(8) pfu VSV and bivalent scFv-dHLX were protected against lethal disease; however, mice infected with immune complexes of 10(8) pfu VSV and monovalent scFv were not. Although repeated scFv-dHLX treatment reduced virus quantities in the blood, neither SCID nor IFN-alphabeta R-/- mice were protected against lethal disease after passive immunization and subsequent VSV infection. This was due to the short half-life of 17 min of scFv-dHLX in the circulation. These data demonstrate that neutralization of VSV and protection against lethal disease do not require Fc-mediated mechanisms and that cross-linking is not crucial for protection against physiologically relevant virus doses in vivo.

Virus validation procedures

The Ad Hoc Working Party on Biotechnology and Pharmacy of the Committee for Proprietary Medicinal Products has developed a number of guidance notes for manufacturers intending to submit applications for market authorization in the European Union which includes one dealing specifically with the validation of production processes for the removal or inactivation of viruses. The strategies used to minimize the risk of viral transmission by biological products are: screening the source materials for viral contamination; examining the ability of the production process to remove or inactivate viruses; and examining the final product for evidence of viral contamination. It is concluded that validation studies provide a significant assurance of viral safety when properly carried out, using appropriate relevant and model viruses.

CTL Escape Viral Variants: I. Generation and Molecular Characterization

Cytotoxic T lymphocytes (CTL) play a pivotal role in preventing persistent viral infections and aborting acute infections. H-2D b-restricted CTL optimally recognize a specific peptide of 9 to 11 amino acids (aa) derived from a viral protein and held in place (restricted) by a MHC class I glycoprotein on the surfaces of infected cells. Only three peptide sequences with the appropriate D b motif from lymphocytic choriomeningitis virus Armstrong strain (LCMV) are known to be presented to CTL by H-2D b molecules; they are from the glycoproteins (GP), residues 33-41 KAVYN FATC (GP1) and 276-286 SGVENPGGYCL (GP2), and the nucleoprotein (NP), 396-404 FQPQNGQFI. Incubation of virally infected H-2 b cells with CTL clones that recognize only GP1, GP2, or NP leads to the selection of viral variants which upon infecting cells hearing H-2 b molecules, escape recognition by CTL of the appropriate specificity. Nucleic acid sequencing showed a single mutation in GP1 (aa 38 F→L), GP2 (aa 282 G→D), or NP (aa 403 F→L) in the variant viruses. When wild-type (wt) LCMV peptides and the three variant peptides (GP1, GP2, NP) were synthesized and subjected to a competitive inhibition binding assay, no differences in binding affinity for H-2D b were found between the wt and variant peptides. Uninfected cells coated with the wt peptide were recognized and lysed by the appropriate CTL clone or by in vivo-primed bulk CTL, but similar targets coated with the GP1, GP2, or NP variant peptides were not. This result, coupled with computer graphic analysis of these variant peptides with the recently solved three-dimensional structure for the D b MHC class I molecule, placed the side chain of the mutated residues on the outer surface of the MHC-peptide complex and accessible to the T cell receptor, Ala substitution at GP residue 38 or 282 or at NP 403 also abrogated CTL recognition and lysis. Inoculation of any one of the mutated viral variants into mice produced an effective CTL response to the other two nonmutated GP or NP peptides, suggesting that production of biologically relevant CTL escape virus variants in vivo requires selection of mutations in more than one and likely all the CTL epitopes, a low probability event.

Viral safety of solvent/detergent-treated blood products.

Laboratory research that began in 1982 led to the licensing in the USA of a solvent/detergent (SD)-treated factor VIII concentrate in 1985. The licence was granted on the basis of several factors. First, studies had demonstrated the inactivation of several marker viruses (vesicular stomatitis virus, Sindbis virus, Sendai virus) and other viruses such as human immunodeficiency virus (HIV), hepatitis B virus (HBV), and non-A, non-B hepatitis virus (NANBHV; now known principally to be hepatitis C virus) added to the factor VIII concentrate just before treatment. Secondly, it had been realized that the relevant viruses in transfusion (e.g. HIV, HBV, NANBHV) all had lipid envelopes. Finally, laboratory, preclinical and clinical evidence indicated that factor VIII and other proteins present in the preparation were unaffected by SD treatment. The applicability of the SD method to a wide range of products and preparations, high process recoveries and a growing body of viral safety information linked with the failure of several other virus-inactivation methods to eliminate hepatitis transmission fostered the adoption of SD technology by more than 50 organizations worldwide. SD mixtures are now used in the preparation of a diverse array of products. Numerous laboratory and clinical studies suggest that coagulation-factor concentrates and other SD-treated products prepared from plasma pools are now safer than the individual units from which they were derived. Also, a large body of evidence indicates that hepatitis A virus (HAV) is not typically transmitted by blood and blood products.(ABSTRACT TRUNCATED AT 250 WORDS)

Virus validation of plasma-derived products produced by Pharmacia, with particular reference to immunoglobulins.

Important factors to assure the safety of plasma-derived products manufactured on an industrial scale are initial screening of the source material and validation of the manufacturing process in accordance with issued EEC guidelines and US [Points to Consider'. Pharmacia's manufacturing process for immunoglobulins contains a specific virucidal step, in which lipid-enveloped viruses are effectively inactivated with a solvent/detergent (SD) combination consisting of 0.3% tri(n-butyl)phosphate and 1% Tween 80. Results from virus validation studies of scaled-down versions of Pharmacia's manufacturing process for immunoglobulins demonstrated extensive removal of relevant and model viruses. More than 5.0 logs of human immunodeficiency virus type 1 (HIV-1) were inactivated in the SD step and, in total, more than 31 logs of HIV-1 were eliminated in the steps studied. Comparison between SD treatment and heating at 60 degrees C of lipid-enveloped viruses in different protein solutions demonstrated that SD treatment is the superior procedure. Polio virus is a model often used in virus validation studies to predict effects on non-enveloped viruses. Because polio virus is more sensitive to heat than are hepatitis A virus (HAV) and human parvovirus B19, thermal inactivation studies with polio virus may result in an overestimation of the effects on HAV and B19.

Molecular characterization of human immunodeficiency virus type 1 cloned directly from uncultured human brain tissue: identification of replication-competent and -defective viral genomes

All presently available replication-competent proviral clones of human immunodeficiency virus type 1 (HIV-1) are derived from cell culture-amplified virus. Since tissue culture is highly selective for viral strains with an in vitro growth advantage, such clones may not be representative of the biologically relevant virus present in vivo. In this study, we report the molecular cloning and genotypic characterization of 10 HIV-1 genomes directly from uncultured brain tissue of a patient with AIDS dementia complex. Targeting unintegrated circular HIV-1 molecules for recombinant lambda phage cloning, we obtained four full-length genomes with one or two long terminal repeats (LTRs), three defective genomes with internal deletions, two rearranged genomes with inverted LTR sequences, and one integrated proviral half with flanking cellular sequences. Nucleotide sequence analysis of these clones demonstrated chromosomal integration, circle formation, genomic inversion, and LTR-mediated autointegration of HIV-1 genomes in vivo. Comparison of a 510-bp hypervariable envelope region among 8 lambda phage-derived and 12 polymerase chain reaction-derived clones from the same brain specimen identified a predominant viral form as well as genetically divergent variants. Variability among 19 of 20 clones ranged between 0.2 and 1.2%. One clone exhibited 8.2% nucleotide sequence differences consisting almost exclusively of G-to-A changes. Transfection of the four full-length HIV-1 genomes identified one clone (YU-2) as replication competent and exhibiting growth characteristics similar to those of tissue culture-derived macrophage tropic strains of HIV-1. These results demonstrate, for the first time, that replication-competent HIV-1 genomes, complex mixtures of defective viral forms, and chromosomally integrated provirus persist in vivo. In addition, the brain-derived viral clones are expected to prove valuable for future studies of macrophage and neurotropism as well as for the analysis of other viral properties that are subject to in vitro selection pressures.

Sequence of the Gene Encoding the Major Neutralization Antigen (VP7) of Serotype 10 Rotavirus

The sequence of the gene encoding the major neutralization antigen (VP7) of the type member (bovine virus strain B223) of the proposed serotype 10 of rotavirus has been determined. This was done using a rapid new strategy involving direct sequencing by primer extension of cDNA generated from the relevant virus gene (gene 8) using a combined reverse transcription/polymerase chain reaction. The sequence obtained is 1062 bp in length and contains a single long open reading frame capable of encoding a protein of 326 amino acids. Comparison of the sequence to that of the corresponding gene of the other major virus serotype found in cattle, serotype 6, showed that it was approximately 25% divergent at the nucleotide level and 18% divergent at the amino acid level.

Inactivation of hepatitis viruses and HIV in plasma and plasma derivatives by treatment with beta-propiolactone/UV irradiation.

A combined treatment of plasma or plasma derivatives by beta-propiolactone (beta-PL)/UV irradiation is in use at Biotest for the preparation of the virus-safe, stabilized serum (Biseko) and coagulation factor concentrates. The efficacy of this sterilization procedure has been demonstrated for HAV (greater than 8.2 log10), HBV (6.9 log10), NANBHV (greater than 4.5 log10) and HIV (greater than 6.0 log10). The methods used in these studies (titration in chimpanzees or cell cultures) are not applicable in routine monitoring of sterilization processes. We therefore developed a test system using four types of bacteriophages: phi X174, phi e, Kappa and f2. Using these bacteriophages in 88 single tests, sterilization efficacy was regularly monitored during the period from 1981 to 1986. The four types of bacteriophages showed, on average, an inactivation rate of 6.7 log10, independent of size or genome structure. This inactivation is in the range of the inactivation of the relevant pathogenic virus, HBV, by beta-PL/UV. It was shown that under the production conditions of Intraglobin and the other Biotest immunoglobulin preparations, beta-PL (without UV) is as virucidal as the combination of beta-PL/UV in plasma or cryo-poor plasma.

Enzymatic amplification of HTLV-I viral sequences from peripheral blood mononuclear cells and infected tissues

Human T-cell lymphotropic virus type I (HTLV-I) and human T-cell lymphotropic virus type II (HTLV-II) have been associated with adult T- cell leukemia/lymphoma (ATL) and a rare T-cell variant of hairy cell leukemia, respectively. Direct detection of viral nucleic acid in peripheral blood lymphocytes (PBLs) and infected tissues in carrier patients and those with chronic disease has proven refractory due to viral transcriptional dormancy and the small number of infected cells present. The investigators report here the successful application of the DNA amplification procedure, termed PCR, to the detection of these human oncoviruses. Judicious selection of specific oligonucleotides for primers and probes provides type-specific and simultaneous detection of these two retroviruses. The ability to amplify and detect highly conserved regions of these medically relevant viruses may facilitate the identification of, as yet, uncharacterized retroviruses.

Radiation sterilization of surgical instruments with a consideration of metal shielding on sterilization efficiency

The feasibility of the use of radiation for sterilization of surgical instruments was evaluated. Two aspects were considered: (1) radiation biology of relevant microorganisms, that is, bacterial spores and viruses, and (2) shielding and radiation protection by the metal of the instruments. After proper cleaning and hot water machine washing, surgical instruments carry few, if any contaminants; however, subsequent handling increases the contamination load. Although large instruments may attenuate as much as 30% of the incident radiation, spores dried on the metal are sensitized to irradiation by some 40%. A dose of 25 kGy (2.5 Mrad) is adequate to inactivate a potential contamination load of approximately 10 7 bacterial spores or approximately 10 4 viruses. Therefore, 25 kGy will provide a high sterility assurance level, and can be recommended with a considerable degree of confidence for hospital-based sterilization of surgical instruments.

Oncogenic Viruses of Domestic Animals

The objective of this article is to review the relevant oncogenic viruses of small animals. Basic scientific principles that have been discovered by research into feline leukemia and other animal cancer viruses have enhanced our understanding of oncogenesis and have led to practical methods of cancer control and prophylaxis.

Generation of hybridomas producing human monoclonal antibodies against herpes simplex virus after [formula omitted] stimulation

Hybridomas producing human monoclonal antibodies against herpes simplex virus were generated by in vitro antigen stimulation before cell fusion. The cell fusion with tonsillar lymphocytes which were stimulated with antigen and/or pokeweed mitogen generated many hybridomas producing human IgG against the virus. A combination of antigen and pokeweed mitogen synergistically enhanced the generation of virus-specific hybridomas. Furthermore, the higher the antibody response of the tonsil, the more virus-specific hybridomas were generated by the cell fusion. These results suggest that cell fusion with in vitro stimulated lymphocytes can be applied to a variety of clinically relevant viruses.

VIRUSES, NEURODEVELOPMENTAL DISORDER AND CHILDHOOD PSYCHOSIS

Four cases are described of prepubertal boys in whom the convergence of neurodevelopmental disorder, viral infection and psychosis seemed more than coincidental. Review of the literature highlights the possibility that viral infection of the central nervous system may play a contributory role in childhood psychosis. Whilst it is essential to avoid a reductionist stance when investigating these difficult conditions, the emergence of potent anti-viral treatments and sophisticated methods of identifying the presence of viral infection should encourage us to consider more carefully the relevance of viruses in childhood psychosis.

Genetics of Resistance to Virus-Induced Leukemias

This chapter discusses the genetics of resistance to virus-induced leukemias. It discusses the genetics of susceptibility to leukemia in mice, the relevance of viruses as etiological agents, and the relationship of this information to leukemias and lymphomas in man. The studies that have formed the foundations of the current knowledge were begun several decades ago, with the period prior to 1960 being characterized by the gradual acceptance of the fact that viruses could cause cancer. The chapter highlights the virology of leukemia-inducing and related viruses, discusses the issues of specificity in virus-host genomic interaction, and describes the current knowledge of host genes conferring resistance or susceptibility to virus-induced neoplasia. The proposition that leukemias are virus induced has been suggested in the studies of the infectious transfer of leukemias in chickens and mice.