ABSTRACT
Studies of the primary cultures of granulocytes, mononuclear, and monocyte-macrophage cells derived from human blood were performed using variola virus (VARV) in the doses of 0.001-0.021 PFU/cell (plaques-forming units per cell). Positive dynamics of the virus accumulation was observed only in the monocyte-macrophages with maximum values of virus concentration (5.0-5.5 Ig PFU/ml) mainly within six days after the infection. The fact of VARV replication in the monocyte-macrophages was confirmed by the data of electron microscopy. At the same time, virus vaccines when tested in doses 3.3 and 4.2 Ig PFU/ml did not show the ability to reproduce in these human cells. The people sensitivity to VARV as assessed from the data obtained on human monocyte-macrophages corresponded to -1 PFU (taking into account the smooth interaction of the virus in the body to the cells of this type), which is consistent to previously found theoretical data on the virus sensitivity. The human susceptibility to VARV assessed experimentally can be used to predict the adequacy of developed smallpox models (in vivo) based on susceptible animals. This is necessary for reliable assessment of the efficiency of development of drugs for treatment and prophylaxis of the smallpox.
Subject(s)
Macrophages/virology , Smallpox/prevention & control , Variola virus/physiology , Virion/growth & development , Adult , Animals , Antibodies, Viral/blood , Granulocytes/immunology , Humans , Macrophages/ultrastructure , Male , Microscopy, Electron , Organ Specificity , Primary Cell Culture , Smallpox/blood , Smallpox/immunology , Smallpox/virology , Smallpox Vaccine/pharmacology , Variola virus/ultrastructure , Virion/ultrastructure , Virus ReplicationABSTRACT
Smallpox disease has been eradicated from the human population since 1979, but is again a concern because of its potential use as an agent of bioterrorism or biowarfare. World Health Organization-sanctioned repositories of infectious Variola virus are known to occur in both Russia and the United States, but many believe other undeclared and unregulated sources of the virus could exist. Thus, validation of improved methods for definitive identification of smallpox virus in diagnostic specimens is urgently needed. In this paper, we describe the discovery of suspected Variola infected human tissue, fixed and preserved for decades in largely unknown solutions, and the use of routine histology, electron microscopy, and ultimately DNA extraction and fluorogenic 5' nuclease (TaqMan) assays for its identification and confirmation.
Subject(s)
Smallpox/diagnosis , Tissue Fixation , Variola virus/isolation & purification , Archives , Bacteriological Techniques , DNA, Viral/analysis , DNA, Viral/genetics , Fluorescent Dyes , Humans , Polymerase Chain Reaction , Sequence Analysis, DNA , Skin/pathology , Skin/virology , Smallpox/virology , Taq Polymerase/genetics , Taq Polymerase/metabolism , Variola virus/genetics , Variola virus/ultrastructureABSTRACT
With increased threat of terrorism, much attention is being directed toward readiness for biodefense. Smallpox virus, a deadly and much feared organism, is among possible bioterrorism agents. Herpesviruses, such as the one that causes chickenpox and shingles, produce skin lesions that may resemble those seen early in smallpox infection. Electron microscopy (EM) is a rapid and reliable method for differentiating poxviruses from herpesviruses. However, before becoming involved in the monitoring of potential smallpox cases, a laboratory must consider several issues, including expertise in virus identification, capacity for handling biohazards, and health and immune status of laboratory staff.
Subject(s)
Bioterrorism/prevention & control , Disaster Planning/methods , Herpesviridae Infections/diagnosis , Herpesviridae/isolation & purification , Smallpox/diagnosis , Variola virus/isolation & purification , Herpesviridae/ultrastructure , Herpesviridae Infections/prevention & control , Herpesviridae Infections/transmission , Microscopy, Electron/methods , Smallpox/prevention & control , Smallpox/transmission , Variola virus/ultrastructureABSTRACT
In order to establish the diagnosis of viral diseases with clinical manifestations in the skin, frequently the help of a virus laboratory is needed. Not even classical childhood diseases such as measles and rubella can safely be diagnosed by the clinician. Today, in many cases diagnosis is possible within hours after the arrival of clinical specimens in the laboratory. Only in cases of enterovirus infection and tropical diseases has the virologist to resort to the classical time-consuming methods of direct (virus isolation) or indirect (serologic) diagnostic virology.
Subject(s)
Skin Diseases/diagnosis , Virus Diseases/diagnosis , Antibodies, Viral/analysis , Dengue/diagnosis , Herpes Simplex/microbiology , Humans , Variola virus/ultrastructure , Warts/microbiologyABSTRACT
A possible use of papain and trypsin immobilized on Biogel P-100 for purification of DNA-containing sheep smallpox viruses was studied. It was shown that the enzymes depolymerize tissue proteins, resulting in a 90% purification of the viruses without causing any considerable morphological changes in the virion structure. It was concluded that fixed proteinases can be effectively used at the final stage of purification of tissue virus-containing suspensions.
