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Stationary tracheal organ cultures of pheasants and chick embryos, treated with a mucolytic agent were used to study the ciliostatic effect of the following viruses: fowl pest (FPV), Newcastle disease (NDV), infections laryngotracheitis (ILV), and infections bronchitis (IBV) - strain Beaudette. In chick embryo tracheal cultures ciliostasis was found to set in as follows: for FPV - at the 24th hour; for NDV - at the 72nd-120th hour; for ILV - at the 168-192nd hour; and for IBV-at the 72nd-96th hour. The ciliostatic effect produced by NDV and IBV coincided in terms of time and dynamics. In pheasant tracheal organ cultures ciliostasis set in as follows: for FPV - at the 24th hour; for NDV - at the 72nd-120th hour; and for ILV - at the 168-192nd hour. This system was shown to be unsusceptible to IBV which produced no effect whatever. In mixed organic cultures, containing several tracheal segments of pheasants and chick embryos each per petri dish IBV did bring about ciliostasis in the chick embryo segments only, within the period after inoculation as cited above. It is stated that pheasant tracheal organ cultures could be employed in the case of an express initial differentiation of FPV, NDV, and ILV, recording the time period for the setting in of full ciliostasis following the inoculation of the respective viruses. Discussed is the possibility of identifying IBV, using mixed tracheal organ cultures of pheasants and chick embryos.
A 29 nm non-cultivable virus (NCV) was detected in faecal extracts from children hospitalized for gastroenteritis. The NCV had a density of 1.35 g/ml in glycerol-potassium tartrate density gradients and was resistant to degradation by proteolytic enzymes, non-ionic detergents and pH extremes. The surface of these virus particles had knob-like projections which appeared to have a symmetrical arrangement. When heated to 56 degrees C, the virus was completely degraded to soluble components which could not be seen by electron microscopy.
Studies have been carried out since 1959 at the Coonoor Influenza Centre to devise a method of cultivating influenza virus in vitro which would be suitable for large-scale virus production. The authors report the successful cultivation of the virus in tissue cultures of chorioallantoic membrane on glass wool. The method described may be used equally satisfactorily for culture in volumes ranging from 1.0 ml to 350 ml, and is as sensitive as cultivation in eggs for the titration of different strains of influenza virus and their neutralizing antibodies. Relatively pure virus for vaccine production and complement-fixing antigen for diagnostic purposes can be produced in large volumes with ease and economy.
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The methylester of griseochelin (1) is a new chemically-made antiviral derivate of the antibiotic griseochelin isolated from fermentations of Streptomyces griseus. It belongs to the polyether group and possesses antiviral activity against enveloped RNA and DNA viruses cultivated in chicken embryo cells (CEC), namely influenzavirus A/WSN, vesicularstomatitis virus (Indiana), vaccinia virus (Lister) and herpes simplex hominis virus type 1 (Kupka). The methylester of griseochelin failed to show virucidal effects on extracellular influenza vacciniavirus particles or to influence virus adsorption and penetration processes. The antibiotic in concentrations of 125-15 micrograms/ml inhibited the virus-induced cytopathic effect of the above mentioned viruses and caused over 90 per cent plaque reduction. Addition of 1 during a one-step growth cycle of influenzavirus A at 4 and 6 h p.i. resulted in complete suppression of virus multiplication at the control niveau of the virus yield accumulated to the same time point. A partial reversibility of the antiviral action against influenzavirus A could be achieved. Coxsackie A9 virus growth in human fibroblast cells was not affected by the inhibitor. Electron-optical observations showed a failure of the formation of the viral capside proteins of HSV type 1 at the second halftime of the replication cycle in CEC-infected and 1-treated cultures.
Human pathogenic viruses can be detected in the hospital environment, on contaminated surfaces or medical instruments. Their transmission to patients or staff has already been reported. Lipophilic viruses (HIV, HBV, HCV,...) are susceptible to many liquid chemicals, but they can survive during short time on inadequately disinfected surfaces. Hydrophilic viruses, without envelope, are more resistant, but generally not associated with severe illnesses. Viruses survival in environment depends on many factors and is always improved with viral aggregation and low temperature, whereas organic matters and relative humidity effects are contrasted. The mechanism of virucide disinfectants is not yet well established, and their targets are not known with precision. Different disinfection procedures (disinfectant concentration, contact time, temperature, pH) can provide a similar virucidal activity on a given virus. The virucidal activity of a disinfectant is evaluated with a cell culture assay in Afnor guidelines. But, there are three major problems with this method, concerning need of high viruses titers, residual disinfectant cytotoxicity on cell culture, and non cultivable viruses. Non standardized tests are also described in papers, but their results can generally not be compared. Molecular biology improvements may lead to reproducible and sensitive tests. At present, no general disinfection procedure effective for most of the viruses, without risks for staff or materials, and with an acceptable economic cost can be recommended.
Noroviruses are positive strand RNA viruses that have received increased attention in recent years because their role as etiologic agents in acute gastroenteritis outbreaks is now clearly established. Much has been learned about the epidemiology of these viruses and the extent of genetic diversity among circulating strains. In contrast, progress on understanding the basic mechanisms of virus replication has been far slower due to the inability to cultivate virus in the laboratory. Despite this limitation, significant progress has been made in defining some basic functions of the norovirus proteins, and the structures of two have been solved to near atomic resolution. This minireview summarizes these recent advances in understanding the structure and function of the norovirus proteins and provides speculation about what roles they may play in the virus replication cycle.
Human noroviruses (NoVs) are the leading cause of food- and waterborne outbreaks of acute nonbacterial gastroenteritis worldwide. As a result of the lack of a mammalian cell culture model for these viruses, studies on persistence, inactivation, and transmission have been limited to cultivable viruses, including feline calicivirus (FCV). Recently, reports of the successful cell culture of murine norovirus 1 (MNV-1) have provided investigators with an alternative surrogate for human NoVs. In this study, we compared the inactivation profiles of MNV-1 to FCV in an effort to establish the relevance of MNV-1 as a surrogate virus. Specifically, we evaluated (i) stability upon exposure to pH extremes; (ii) stability upon exposure to organic solvents; (iii) thermal inactivation; and (iv) surface persistence under wet and dry conditions. MNV-1 was stable across the entire pH range tested (pH 2 to 10) with less than 1 log reduction in infectivity at pH 2, whereas FCV was inactivated rapidly at pH values < 3 and > 9. FCV was more stable than MNV-1 at 56 degrees C, but both viruses exhibited similar inactivation at 63 and 72 degrees C. Long-term persistence of both viruses suspended in a fecal matrix and inoculated onto stainless steel coupons were similar at 4 degrees C, but at room temperature in solution, MNV-1 was more stable than FCV. The genetic relatedness of MNV-1 to human NoVs combined with its ability to survive under gastric pH levels makes this virus a promising and relevant surrogate for studying environmental survival of human NoVs.
The photodynamic effect of toluidine blue (TB) was studied on MM virus cultivated in L cells and on the RNA extracted form MM virus inoculated to mice by intracerebral route. MM virus infectivity is completely inhibited by the photodynamic action of TB, due to lesions at the molecular level of the viral RNA.
Human immunodeficiency virus (HIV) was cultivated directly from whole blood treated with anticoagulant by cocultivation with phytohaemagglutinin-stimulated cord blood lymphocytes. When heparin was used as the anticoagulant, isolation rates were low (10% to 56%, depending on the patient group); but when EDTA was used, isolation rates were much higher (50% to 100%). Culture of whole blood gave results identical to those of culture of separated peripheral mononuclear cells, and in some cases virus could be isolated from as little as 10 microliters of unseparated EDTA anticoagulated blood.
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Sendai virus multiplication in the presence of ceruloplasmin resulted in the appearance of qualitatively modified progens. There was an increase in the proportion of incomplete virus particles and a decrease in hemagglutinin and neuraminidase activities, in virus infectivity and antigenicity (with the exception of subviral fractions with predominant hemagglutinating activity). Virus progens obtained in the presence of ceruloplasmin had a higher sensitivity to detergent treatment; virus ghosts formed by re-aggregation of envelope fragments could be observed in the electron microscope.
Influenza A (H1N1) viruses when initially isolated in mammalian cell cultures (MDCK cells) had different agglutination reactions with chicken and guinea-pig erythrocytes compared to the same viruses after passage. On first isolation the virus HA resembled the 'O' phase viruses described originally by Burnet and Bull and agglutinated mammalian but not avian erythrocytes. After passage, the virus HA resembled a classical 'D' phase virus and agglutinated both avian and mammalian erythrocytes. Monoclonal and polyclonal antisera detected antigenic differences between the HAs of the viruses in the 'O' and 'D' phases. The 'O' phase virus HA reacted preferentially with antibodies in post infection human antisera. Viruses in the 'O' phase replicated poorly in the allantoic cavity of embryonated hens' eggs whilst 'D' phase virus replicated in both MDCK cells and in embryonated hens' eggs. At least three distinguishable subpopulations of influenza A (H1N1) viruses may co-exist in clinical throat swab material, including viruses possessing HAs in the 'O' and 'D' phases and other 'D' phase viruses cultivable in embryonated hens' eggs but antigenically distinguishable from the corresponding 'D' phase virus in MDCK cells.
The biotechnologic developments during the last decade have led to the production of inactivated poliovirus vaccine (IPV) on an industrial scale and at economically acceptable costs. Replacement of primary monkey kidney cells by subcultured monkey kidney, Vero, or human diploid cells as substrate for virus multiplication as well as the introduction of the microcarrier culture technique have made cell and virus cultivation in large fermentors of 100-1,000 liters feasible. Procedures for processing virus harvests into highly concentrated purified vaccines were developed; also, the safety and potency control tests were improved and simplified. It has been demonstrated that these more potent poliovirus vaccines, either alone or in combination with diphtheria-tetanus-pertussis vaccine, induce a high immunity with reduced vaccination schedules. The overall costs of vaccination will be reduced considerably in this way. In addition, the results of biochemical and immunologic studies indicate that neutralizing antibodies can be induced by the viral proteins alone. These findings open up promising perspectives for production of subunit poliovirus vaccines with use of recombinant DNA and synthetic antigen, a method that has already proved feasible for producing vaccine against foot and mouth disease. These new techniques may lead to a further reduction of production costs and will improve the safety of the vaccine.
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Streptovirudins are new antibiotics isolated as a mixture of several structurally related compounds from fermentations of Streptomyces griseoflavus (Krainsky) Waksman et Henrici var. thuringensis JA 10124. They possess antiviral activity against RNA and DNA viruses cultivated in chick embryo cells, namely Sindbis, fowl plague, Newcastle disease (NDV), pseudorabies, vaccinia and sheep abortion viruses. The naturally formed streptovirudin complex, in concentrations of 20-2.5 mug/ml inhibited the viral cytopathic effect and caused 100 percent plaque reduction. Mengo, Coxsackie B1-B5, ECHO 30 and 33, and polio (wild and attenuated types 1, 2, and 3) viruses grown in FL cells were not sensitive in the agar-diffusion plaque-inhibition test. The antibiotics failed to show a direct virucidal effect on the NDV virion itself or to influence virus adsorption and penetration processes. Addition of streptovirudin complex during a one-step growth cycle of NDV from 0-4 hours after virus adsorption resulted in complete suppression of virus yield. The antibiotic complex consists of two main groups: I - A1, B1, C1, D1, E1 and II - A2, B2, C2, D2, E2, each of which possess antiviral activity.
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