Beverley T. Perry

Influenza Virus Nucleic Acid: Relationship between Biological Characteristics of the Virus Particle and Properties of the Nucleic Acid.

SUMMARY: Five A strains and three B strains of influenza virus were purified and found to contain ribonucleic acid in amounts varying from 0·75 to 1·1 %. The proportion of the purine and pyrimidine derivatives in the nucleic acid of each strain was determined. When the ratio adenine + uracil: guanine + cytosine was evaluated, the following values were obtained: A strains —PR8, 1·27± 0·02; MEL, 1·22 ± 0·01; WSE, 1·26 ± 0·01; SWINE, 1·24 ± 0·04; CAM, 1·28 ± 0·01. B strains—LEE, 1·42 ± 0·04; MIL, 17·43 ± 0·05; ROB, 1·38 ± 0·01. The nucleic acid content of PR8 virus preparations of varying degrees of incompleteness was determined both by specific absorption at 260 mμ. and by estimation of phosphorus present in the nucleic acid extract. As measured by both methods, virus preparations of low infectivity were found to have a decreased nucleic acid content. The demonstration of specific differences in the nucleic acid of A and B strains and of the relationship between the infectivity and nucleic acid content of the virus affords strong evidence that the nucleic acid is an intrinsic part of the influenza virus particle.

Biological and Physical Properties of the Ryan Strain of Filamentous Influenza Virus.

SUMMARY: An influenza A strain, Ryan, isolated in Melbourne in 1954 contains on a particle basis about 20% filaments. By using the electron microscope for particle counting, the efficiency as haemagglutinin of Ryan ‘long’ filaments (particles in which length: width → 6) was estimated to be about 6 times greater than that of spheres of PR 8 virus. Allantoic fluid preparations of Ryan virus have high values for the ratio (log) EID50:AD and estimates indicate that a maximum of 6 ‘long’ filaments are equivalent to one EID50, compared with 18 particles in the case of PR 8 virus or 14 particles in the case of a mutant strain of Ryan virus which exists almost exclusively as spheres. Compared with virus PR 8 spheres (16 particles: AD), spherical particles present in filamentous Ryan preparations have a low efficiency as haemagglutinin (29 to 38 particles: AD). These particles are formed before the ‘long’ filaments, and in fluids harvested 21 hr. after inoculation when the proportion of ‘long’ filaments is very low, about 9 spherical particles constitute 1 EID 50. For the purpose of subsequent chemical examinations filaments were isolated and concentrated by a process involving specific adsorption to and elution from red cells, followed by 3 cycles of differential centrifugation. The ratio of filaments: spheres in such preparations is about 50:50. ‘Purified’ filaments weigh about 30 times as much as the spherical particles present and account for over 95% of the total weight present in purified preparations.

Properties of the nucleic acid of the Ryan strain of filamentous influenza virus.

SUMMARY: The amount, type and properties of nucleic acid have been estimated in the two end fractions, Ryan supernatant (Ryan Sup.) and Ryan final filaments (Ryan F.F.) of a purification procedure applied to allantoic fluid preparations of filamentary Ryan virus. Ryan Sup. consists of 80% spherical particles and 20% short filaments (length:diameter < 6). Analysis indicates an average nucleic acid content of 0.75% (PR 8 = 0.91%). The value of the ratio, adenine + uracil: guanine + cytosine for the nucleic acid of A strains of influenza virus varies between 1.22 and 1.28. The value for Ryan Sup. is 1.20; for a mutant strain of Ryan virus which exists almost exclusively as spheres, the value is also 1.20. The ratio of filaments to spherical particles in Ryan F.F. is about 50:50. On a dry-weight basis, Ryan F.F. contains about 0.25% RNA. There may be small amounts of DNA present. On a particle basis, the residual filamentary structures contain about eight times as much RNA as do PR 8 spheres. The value of the above ratio of bases however is about 0.9. Treatments of filaments with diethyl ether releases a soluble complement-fixing antigen (CFA) which on purification is found to have the same nucleic acid content per unit of CF activity as does soluble CFA isolated from either PR 8 or Ryan Sup. viruses. In each case the value of the above ratio of bases is about 1.25. On a particle basis filaments in Ryan F.F. contain 3 to 4 times as much soluble CFA as does PR 8 virus so that the residual RNA in Ryan F.F. must closely correspond in properties to RNA from the potential host cell which has a value for the above ratio of about 0.6. Exposure of Ryan F.F. to ribonuclease or to a procedure which degrades the filamentary form to smaller spherical units does not affect the amount or properties of the associated RNA. A tentative scheme is proposed for the formation of virus particles of Ryan F. preparations. It is postulated that most of the spheres present in such preparations arise by fragmentation of the tip of forming filaments where there is a relative concentration of viral type RNA. The filaments which are found in the allantoic fluid thus represent only part of the original filamentary structures. This concept implies that filaments break more readily at those places where there is an enrichment of viral type nucleic acid.