Group-specific Complement-fixing Antigen Research Articles

Characteristics of an adenovirus type 19 conjunctivitis isolate and evidence for a subgroup associated with epidemic conjunctivitis

Although adneovirus type 19 (Ad19) was first described in 1955, this virus was not associated with disease until its isolation from outbreaks of conjunctivitis in 1973. A strain of Ad19 isolated from a case of conjunctivitis in Seattle in 1974 was compared with the reference strain (3911). Plaque number and size were enhanced by 30 mM MgCl2. Low pH and chloroform treatment had no effect on either strain's activity, but the two strains were sensitive to pH 8. Growth curves were characteristic of adenoviruses, but differences were seen in the amount of virus released. The ratios of particles to plaque-forming units (approximately 10,000:1) were similar for both. Both virus preparations contained high concentrations of group-specific complement-fixing antigen. Cross-reactions were seen by hemagglutination inhibition and immunoelectron microscopy between antisera to Ad8, Ad9, and Ad10 versus both strains of Ad19, but were not seen by neutralization. We would like to suggest, based on exclusive conjunctivitis association and cross-reactions, that the four cross-reacting serotypes, Ad8, Ad9, Ad10, and Ad19, represent a subgroup of adenoviruses specfically associated with conjunctivitis.

Origin and structure of the group-specific, complement-fixing antigen of Rickettsia rickettsii.

Rickettsia rickettsii was treated with ether and examined by negative-contrast electron microscopy. Group-specific complement-fixing antigen was seen to be originating from the cell wall. The antigen was composed predominately of round particles 10 to 60 nm in diameter. Intact R. rickettsii and antigen from ether-treated organisms were purified by density gradient centrifugation and analyzed by polyacrylamide gel electrophoresis. The whole rickettsial cell was composed of a minimum of 30 proteins which ranged in molecular weight from about 23,000 to 155,000. The "soluble" antigen contained nine proteins ranging in molecular weight from about 28,000 to 150,000.

Origin and Structure of the Group-Specific, Complement-Fixing Antigen of Rickettsia rickettsii

Rickettsia rickettsii was treated with ether and examined by negative-contrast electron microscopy. Group-specific complement-fixing antigen was seen to be originating from the cell wall. The antigen was composed predominately of round particles 10 to 60 nm in diameter. Intact R. rickettsii and antigen from ether-treated organisms were purified by density gradient centrifugation and analyzed by polyacrylamide gel electrophoresis. The whole rickettsial cell was composed of a minimum of 30 proteins which ranged in molecular weight from about 23,000 to 155,000. The "soluble" antigen contained nine proteins ranging in molecular weight from about 28,000 to 150,000.

In vitro transformation of canine embryo cells by murine sarcoma virus (Kirsten).

SummaryCanine embryo cells were transformed in vitro by Kirsten murine sarcoma virus (Ki-MSV). The transformed cells contained group-specific complement-fixing antigen characteristic of the murine leukemiasarcoma virus complex. Cell-free supernatant fluids from the transformed cultures were infectious for rat, guinea pig, and canine embryo cells. RNA-dependent DNA polymerase activity was demonstrated in the transformed cells. Electron microscope observation showed that the transformed cells produced type C virus particles. The present results indicate that members of the murine sarcomaleukemia virus complex, particularly Ki-MSV, exhibit a broader host range than was hitherto believed.

Long-term in vitro cultivation of Rauscher Leukemia virus in rat embryo cells.

SummaryThe long-term in vitro cultivation of Rauscher leukemia virus (RLV) in rat embryo (RE) cells is reported. RE cells infected initially with RLV have replicated infectious virus and produced complement-fixing (CF) antigen characteristic of the murine leukemia-sarcoma virus complex for more than 12 months. Production of complete virus has been demonstrated by means of electron microscopy; and of infectious virus by the in vitro assay technic (COMuL test) based on detection of the group-specific CF antigen of murine leukemia viruses. Interferon production was not demonstrated in infected cultures. The data indicate that RE cells could be used for cultivation of RLV in a carrier state.

Investigations on virus production in RSV mammalian tumours.

SUMMARY Six different Rous mammalian tumour lines were studied for the occurrence of virus specific antigens, the presence of the viral genome and the synthesis of infectious and non-infectious particles. Four Rous tumour lines were induced in stumice presumably free from latent murine leukemia virus (MLV), three of them by a cloned Schmidt-Ruppin strain of Rous sarcoma virus (sr-RSV) belonging to sub-group D, the fourth with an uncloned sr-RSV strain. In addition a Prague(P)-RSV induced mouse tumour line (RVP3) and an sr-RSV induced hamster tumour line (RSH) and sr-RSV induced chicken tumours were likewise investigated. In all mammalian lines, except for RVP3, avian leukosis group specific complementfixing antigen was found. Tumour specific transplantation antigen was demonstrated in all mammalian lines except RSH, whereas no infectious RSV could be detected in any of the mammalian tumours. Likewise electron microscopy revealed no virus particles in more than 9500 cell sections of RSV-stu tumours, but the viral genome was shown to be present by transfer of the tumour to chickens. In the RVP3 line virus particles were detected resembling typical murine leukemia virus while in the RSH line particles were found morphologically similar to the latent hamster virus described by Bernhard & Tournier (1964). The interpretation and significance of the findings are discussed.

Biophysical comparison of two canine adenoviruses.

The two canine adenoviruses, infectious canine hepatitis (ICH) virus and infectious canine laryngotracheitis (ICL) virus (also designated as Toronto A26/61 virus), were studied with respect to their morphology and the biological properties of their soluble components. The two viruses were found to be composed of soluble components similar to, and carrying biological activities corresponding to, those of the human adenoviruses after fractionation by rate zonal centrifugation and anion-exchange chromatography. The following soluble components were identified: hexons (carrying a common group-specific complement-fixing antigen), penton oligomers (complete soluble hemagglutinin), and penton and fiber monomers (incomplete soluble hemagglutinins). The latter were indicated by hemagglutination enhancement with selected antisera directed against human adenovirus soluble components. Elution characteristics of corresponding components of the two viruses in anion-exchange chromatography experiments were distinctly different. Electron microscopic examination of purified virions and soluble components revealed the fiber component of ICL virus to be 35 to 37 nm in length, and that of ICH virus to be 25 to 27 nm long.

Biological characterization of structural components of Adenovirus type 12.

SUMMARY Two products with a large mass and five different soluble components of adenovirus type 12 have been identified. The former were virus particles and empty capsids, which both carried group-specific complement-fixing (CF) antigen and were active as complete haemagglutinins (HAs). Their buoyant densities in CsCl were 1·330 to 1·340 and 1·295 to 1·305, respectively. The soluble components were identified as hexons, pentons, isolated fibres, dimers of fibres and isolated vertex capsomeres. Hexons were trypsin-resistant, ther-mostable and carried group-specific CF antigen; ultrastructurally they resembled hexons of other human adenoviruses. Pentons were active as an incomplete HA, causing agglutination in the presence of heterologous antisera against representative members of all three subgroups of adenoviruses. Furthermore, they were trypsin-sensitive and thermolabile. Isolated fibres represented another incomplete HA, exhibiting agglutinating activity only in the presence of antisera against members of subgroup III, e.g. type 2. This component was also identified as the dominating type-specific CF antigen. The length of fibres was estimated as 28 to 32 nm. by electron microscopy. The dimers of fibres represented a complete HA, the activity of which became apparent only after other material was removed. Both isolated fibres and their dimers were trypsin-resistant and thermostable. The assumed pentons and fibre dimers occurred in too low concentration to allow ultrastructural identi-fication. Isolated vertex capsomeres were demonstrated by a haemaggluti- nation enhancement antibody consumption test. They were completely destroyed by trypsin treatment and exhibited a moderate thermostability. Electron microscopy confirmed the presence of capsomere-like structures. These structures occasionally displayed a five-sided contour, thereby differing from hexons. The sequence of elution of different components from an anion exchanger was: isolated vertex capsomeres, hexons + pentons, fibres and dimers of fibres. In zonal centrifugation experiments the components sedimented at decreasing rates in the following order: hexons, pentons+isolated vertex capsomeres, dimers of fibres and isolated fibres. Probably on account of the morphological characteristics of the various components they presented a different pattern of elution in exclusion chromatography. Dimers of fibres, pentons and even isolated fibres were eluted before hexons. The position of the peak activity of type 12 fibres in the elution diagram was similar to that of types 1, 2 and 5 fibres.

Separation and characterization of soluble adenovirus type 9 components.

Four different soluble components of adenovirus type 9 (Rosen's group II) were identified. These were a complete hemagglutinin (HA), an incomplete HA, components carrying group-specific complement-fixing (CF) antigen, and components identified only by their hemagglutination-inhibition (HI) antibody consuming capacity and antigen activity in CF tests with an antiserum against complete HA. The complete HA sedimented relatively rapidly. It was composed of 12 pentons (vertex capsomers plus projections) aggregated into the form of a pentagonal dodecahedron. The length of the projections was about 12 to 13 mmu. Thus they appeared longer than the corresponding structures of types 3 and 11, but shorter than those of types 4 and 5. The rate of sedimentation of complete HA of type 9 was intermediate to those of the complete HA of types 3 and 11. The incomplete HA sedimented together with components carrying group-specific CF antigen, but could be separated from those by anion-exchange chromatography. Two different antigens were present in incomplete HA. One could absorb a group-specific hemagglutination-enhancing antibody, and was sensitive to treatment with trypsin. The other antigen could absorb the type-specific HI antibody and was not destroyed by trypsin. In addition to the incomplete HA, a separate population of more slowly sedimenting components showed a capacity to absorb HI antibody. These components could also be identified in CF tests when an antiserum against complete HA was applied. The incomplete HA, group-specific CF antigen, and slowly sedimenting HI antibody absorbing components are suggested to represent isolated penton, hexon, and fiber components, respectively.

The relationship between the soluble antigens and the virion of adenovirus type 3: I. Morphological characteristics

The soluble antigens of adenovirus type 3 were purified by zone centrifugation. The type-specific antigen, the hemagglutinin (HA), sedimented as a homogeneous population of particles at a rate corresponding to 50–60 S. Morphologically the HA appeared as six- or five-pointed stars with an overall diameter of 40–50 mμ. These appeared to be composed of a number of identical, tubular, capsomere-like structures, each associated with a thin club-shaped projection. The complete HA was interpreted as being formed out of 12 such capsomere-like structures arranged in accordance with icosahedral symmetry, each of them representing one facet in a pentagonal dodecahedron, and the club-shaped projections extending radially. The capsomere-like structure had an inner diameter of 20–25 A, outer diameter of 60–85 A, and was 55–80 A long, thus being somewhat smaller than the majority of free-lying capsomeres. The club-shaped projections had a length of 80–110 A and a maximum width of 40–60 A. Similar projections were found extending from the vertices of purified intact virions. The HA therefore is proposed to be a strictly symmetrical aggregate of that type of morphological subunits, which can also form the vertices of the intact virion. The particles carrying group-specific complement-fixing antigen activity had a sedimentation constant of 13 S. Their ultrastructure was identical with that of the majority of free-lying capsomeres as seen released by spontaneous disrupture of purified intact virions. The dimensions of the tubular structures were the following: inner diameter 20–30 A, outer diameter 80–90 A, and length 70–80 A.

Virus particles associated with “nonproducer” Rous sarcoma cells

Examination of four cultures of “nonproducer” (NP) Rous sarcoma cells in the electron microscope revealed the presence of extracellular particles with the morphology of avian leukosis viruses. Focus assays on fluids from these NP cell cultures failed to detect infectious virus. To confirm this observation, two lines of NP cells were carried in culture through seven serial passages and examined at intervals for evidence of infectious or noninfectious virus. A total of 160 ml of culture fluid was removed from the two NP cell lines at various times during passage and this fluid was tested for infectious virus in fresh cultures of chick fibroblasts. No evidence of infection was found when the indicator cells were tested by focus assay, RIF test, or COFAL test, and the indicator cells remained free of virus particles detectable in the electron microscope. NP cells were examined directly for the presence of group-specific complement-fixing antigen with hamster antiserum against Schmidt-Ruppin sarcoma virus. In early passages no antigen was detected, but the 6th and 7th passage NP cells contained small amounts of C'F antigen. Electron microscopic examination revealed the presence of virus particles in both cell lines through 7 passages in culture. Direct particle counts were not attempted; however, NP cell cultures were estimated to contain 1 10 to 1 100 the number of particles seen in cultures of Rous sarcoma cells that were releasing infectious virus.