[The mechanisms of the antiviral action of the bora-adamantane derivative preparation BG-12]. / Izuchenie mekhanizmov protivovirusnogo deistviia proizvodnogo boraadamantana preparata BG-12.

The effect of one of the derivatives of boraadamantane, preparation BG-12, on reproduction of influenza type A and B viruses was studied. This preparation was shown to inhibit multiplication of a wide range of influenza type A and B virus strains. It is important that BG-12 inhibits in cell culture the replication of a mutant of fowl plague virus A/FPV/Weibridge resistant to remantadine. BG-12 was found to exert a marked viricidal effect, to inhibit the hemolytic activity of both influenza A and influenza B viruses, but not the neuraminidase activity of these viruses. The synthesis of some virus-specific proteins was found to be disturbed in influenza B/Ann Arbor/86-infected cells in the presence of BG-12. A FPVBAR mutant of influenza A/FPV/Weibridge (H7N7) virus resistant to BG-12 was obtained. All the BG-12-resistant reassortants inherited genes 4 and 7 coding for hemagglutinin (HA) and membrane proteins (M) from FPVBAR mutant, respectively, and the remaining genes from BG-12-sensitive A/Krasnodar/101/59 (H2N2) virus. The sequencing of a region of the M gene encoding for the transmembrane protein M2 revealed the substitution of Ala30-Thre in this protein. Multiple attempts at generating a mutant of influenza type B virus resistant to BG-12 failed.

[Factors that cause a change in the antigenic structure of of the influenza virus hemagglutinin]. / Faktory, vyzyvaiushchie izmenenie antigennoi struktury gemaggliutinina viursa grippa.

Crossing of norakin-resistant mutant NR1 of A/Waybridge (H7N7) strain of fowl plague virus (FPV) with human influenza virus strains produced recombinants inheriting the hemagglutinin (HA) gene of the NR1 mutant and neuraminidase (NA) genes of human influenza virus strains. The R120 recombinant produced by crossing of NR1 with A/Taiwan/1/86 (H1N1) strain, unlike other recombinants and NR1 mutant, lost the capacity of reacting in H1 test with two monoclonal antibodies (MCA) to HA7: 71/4 and 46/6. The ts mutant A/FPV/Rostok which has ts-mutation in HA-gene also had changes in the antigenic specificity of HA. The RA and RB recombinants produced by crossing R120 with the A/Krasnodar/101/59 strain and inheriting HA-gene from R120 and NA-gene from A/Krasnodar/101/59 strain recovered the initial HA antigenic structure. No changes in the antigenic properties of HA were observed in the recombinants produced by crossing the original A/FPV/Waybridge strain with A/Taiwan/1/86 strain and inheriting HA-gene from the original A/FPV/Waybridge strain and NA-gene from A/Taiwan/1/86 strain. It is concluded that ts mutations in influenza virus HA-gene may be accompanied by changes in the antigenic specificity of this virus HA. The possibilities of manifestation of phenotypic suppression at the level of influenza virus virion membrane proteins and the causes of changes in the HA antigenic structure in this virus recombinants are discussed.

Relation between drug resistance and antigenicity among norakin-resistant mutants of influenza A (fowl plague) virus.

Norakin-resistant (NR) mutants of fowl plague virus (A/FPV/Weybridge, H7N7) have 1 to 2 (in one instance 3) amino acid substitutions in different positions of the heavy (HA 1) and/or light (HA 2) subunits of the haemagglutinin (HA) molecule. Investigation of NR mutants using the haemagglutination inhibition test with monoclonal antibodies (MAb) to the HA of A/seal/Massachusetts/80 (H7N7) virus revealed that one of the mutants (NR 1) differs antigenically from the wild-type fowl plague virus: its haemagglutination was not inhibited by MAb 55/2 and 58/6. By contrast, MAb-resistant (escape) mutants, selected from the wild-type fowl plague virus under pressure from MAb 55/2 or 58/6, showed reduced drug sensitivity. These findings suggest a possibility of correlation between alteration of influenza virus antigenicity and change of its sensitivity to drugs whose target is the haemagglutinin. This potential effect should be taken into account when antiviral substances directed to surface influenza virus antigens are being developed for use as antiviral drugs.

Correlation of amino acid residues in the M1 and M2 proteins of influenza virus with high yielding properties.

The ability of influenza A viruses to replicate to high titer in the allantoic sac of the chicken embryo has been mapped to the matrix protein gene (RNA 7). Because influenza A/WSN/33 (H1N1) virus grows poorly in this host but contains a matrix protein gene with a sequence similar to sequences from viruses that grow well in eggs, we derived a single gene reassortant containing only the M gene from A/WSN/33 (H1N1) in a background of the other 7 RNA segments from A/Philippines/2/82 (H3N2) (a low yielding virus, hy-). This reassortant replicated 10 times better than the A/WSN parent itself, indicating that the high yielding (hy+) phenotype of the A/WSN/33 M gene may be suppressed by one of the other genes of A/WSN/33. Comparison of M gene sequences between hy+ (including A/WSN/33) and hy- strains allowed us to correlate specific amino acid positions in M1 and M2 proteins with the growth properties of influenza viruses.

Reproduction of human and animal influenza viruses in human nasal polyp organ cultures.

Human influenza virus strains were easily grown and passaged in human nasal polyp organ cultures causing marked damage of the epithelium. Unlike to human strains, the animal influenza virus strain could be propagated for no longer than 2 or 3 passages and even the 1st passage failed to cause significant morphological changes of the epithelium cells.

Studies of a recombinant which inherited the haemagglutinin from the human influenza virus A/Hong Kong/1/68 (H3n2) and other genes from influenza virus A/duck/Ukraine/1/63 (H3N8).

A recombinant (R3/3) has been obtained which inherited the gene coding for the haemagglutinin from human influenza virus A/Hong Kong/1/68 (H3N2), and seven other genes from influenza virus A/duck/Ukraine/1/63 (H3N8). The recombinant R3/3 had properties, a ts+ phenotype and a capability of reproducing in chick embryos, similar to those of the duck influenza virus, but, in contrast to this parent, had lost its ability to reproduce in organ cultures of colon of ducks and chickens as well as in monolayer cultures of chick embryo fibroblasts.

Studies on a temperature-sensitive mutant of fowl plague virus having a mutation in gene 7 coding for the M protein.

A fowl plague virus (FPV) temperature-sensitive mutant, ts 303/1 having a ts mutation in gene 7 coding for the matrix (M) protein has been obtained. The mutant induced synthesis of virus-specific RNA and polypeptides as well as ribonuclear protein (RNP) formation in cells under non-permissive conditions; however, haemagglutinin cleavage was reduced, functionally active haemagglutinin and neuraminidase were absent and virions were not formed. In mutant-infected cells at 36 degrees C haemagglutinin cleavage was also reduced and virions formed had an altered NP:M ratio as well as a decreased haemagglutinin content. A population of virions formed under these conditions was heterogeneous both in morphology and in buoyant density. The data obtained suggest that a mutation in the M proteins of orthomyxoviruses can affect processing of the haemagglutinin and impair final stages of virion morphogenesis.

Studies of biological properties of the recombinants between human influenza and fowl plague viruses as related to genome composition.

Some biological properties of recombinants obtained by crossing of fowl plague and human influenza viruses were studied. The capacity of the recombinants to reproduce in chick embryo fibroblast cultures was in reverse correlation to the number of genes coding for P proteins derived from the human influenza virus. The genome composition was of importance for the expression of ts-phenotype of the recombinants in different systems. Substitution of at least one gene in the fowl plague virus genome by a corresponding human influenza virus gene resulted in the decrease of virulence for 1-day-old chickens. The presence of three P genes from human influenza virus genome in the genome of the recombinant proved to be insufficient for the capability of the recombinant to reproduce in organ cultures of human origin.

The replication of influenza A viruses in organ cultures of human nasal polyps.

Organ cultures of human nasal polyps were shown to support the replication of five out of seven human influenza A viruses and three out of six avian strains with varying degrees of efficiency. The ability to replicate was independent of the antigenic formula of the virus. The structure of nasal polyps closely resembled that of normal nasal mucosa and infection with influenza A virus resulted in histological changes analogous to those seen in natural infections. This system provides an in vitro method for more detailed studies of influenza A virus and possibly other respiratory virus infections of man.

Investigation on the mechanisms of the failure of human influenza virus to replicate in chick embryo cell cultures.

Thirteen strains of human influenza virus producing in chick embryo cell (CEC) cultures either virions with low infectivity or no virions were studied. In CEC, most of the strains induced synthesis of viral RNA, polypeptides, and ribonucleoprotein and produced functionaly active haemagglutinin, neuraminidase and virions lower infectivity. The low infectivity of virions produced by strains of this functional group was due to disturbed cleavage of a polypeptide, haemagglutinin precursor, formed in CEC, into a heavy a light haemagglutinin chain. Two strains belonging to another functional group induced no synthesis of virus-specific macromolecules in CEC, but were able to adsorb onto the cells. With one of these viruses, no transcription of parental RNA could be detected in CEC. There was no relationship between the grouping of the studied strains into a certain functional group with their antigenic characteristics, the year of isolation, the passage history in chick embryos and human pathogenicity.

[Method for preparing monospecific antiserum to fowl plague virus hemagglutinins]. / Metod polucheniia monospetsificheskoi antisyvorotki k gemaggliutininu virusa chumy ptits

A simple method for preparation of monospecific antiserum for the hemagglutinin of fowl plague virus has been developed. It is based on selective inactivation of the enzymatic and antigenic properties of neuraminidase by heating of the virus at 56degrees for 3 hours followed by disruption of the preparation with detergent and removal of the inner proteins by ultracentrifugation. Immunization of animals with such preparations produced antiserum containing considerable amounts of antibody for the hemagglutinin in the absence of antibody for other proteins of fowl plague virus.