Antigenic variation in influenza viruses, with special reference to Hong Kong influenza*

Discussion Papers-Session II Antigenic Variation in Influenza Viruses, with Special Reference to Hong Kong Influenza * by R. G. WEBSTER a The studies on antigenicrelationships ofHongKong influenza virus reported at this Conference b, C, d have shown that " antigenic drift" has occurred in the neuraminidase antigen of A2 influenza viruses since their occurrence in 1957 and has continued through into the Hong Kong influenza virus. The haemagglu- tinin antigen of Hong Kong virus, on the other hand, was antigenically distinct and a major change, or " antigeniQ shift " had occurred in this antigen. c, e The separation of the surface antigens of influenza viruses, either by recombination and segregation of antigens in vivo or by physical means in vitro, has been shown to be important in eliminating steric inter- actions between antigens and antibodies that can give equivocal cross-reactions. The antigenic cross-reactions between the haemag- glutinin of Hong Kong influenza virus and A/Equi-2/ 63, Duck/Ukraine/2/60 and Duck/Ukraine/1/63 f influenza viruses, and the reactions between the neuraminidase of Hong Kong influenza virus and the neuraminidase from 2 different duck influenza viruses d lead us into the question of how influenza viruses of humans containing haemagglutinins or neuraminidase from animal or avian viruses arise. The examples available suggest that these reactions cannot be coincidental; they cannot arise from independent variation in different viruses. Genetic interaction between human and animal or avian strains may play a role in the evolution of pandemic human influenza viruses. Kasel & Couch g have * This work was supported by Public Health Service Grant AI-08831 from the National Institute of Allergy and Infectious Diseases, the Hartford Foundation and ALSAC. a Laboratory of Virology and Immunology, St Jude Children's Research Hospital, 332 N. Lauderdale, Memphis, Tenn. 38101, USA. b See paper by M. T. Coleman & W. R. Dowdle on page 415 of this issue. =c See paper by J. L. Schulman & E. D. Kilbourne on page 425 of this issue. d See paper by G. C. Schild & R. W. Newman on page 437 of this issue. e See paper byW. R. Dowdle et al. on page 419 of this issue. f See paper by B. Tumova & B. C, Easterday on page 429 of this issue. g See paper by J. A. Kasel & R. B. Couch on page 447 of this issue. approached this question and shown that A/Equi-2/ Miami/63 virus will cause infection in humans and that A2/Hong Kong/68 virus of humans will cause infection of Chincoteague ponies. An explanation of the occurrence of human influenza viruses with animal or avian antigens should now be sought in a positive fashion. The tools are at hand for producing antigenic hybrids of human or animal influenza viruses in the laboratory and the next question we should try to answer is whether virulent antigenic hybrids can be produced in natural hosts. Information is also lacking on the importance of the neuraminidase antigen in protection of humans against influenza virus infection. The information presented in this session shows that the human population had been exposed to a virus that contained the same neuraminidase that is present in the Hong Kong influenza virus and yet an epidemic of influenza was caused by Hong Kong virus. It has been shown that antibodies to the neuraminidase of influenza virus are found in the sera of humans after infection ' and that vaccination ofmice with isolated neuramini- dase will cause a reduction in spread after challenge with viruses carrying the homologous neuraminidase irrespective of the haemagglutinin antigen. Several questions arise regarding the Hong Kong epidemic and neuraminidase antibodies: (1) were antineurami- nidase antibodies to A2/Hong Kong influenza detectable before the onset of the epidemic? (2) Is antibody to the neuraminidase " short-lived " ? (3) Was antibody to neuraminidase present in human nasal secretions? The results in Table I show that children bled before the'onset of the Hong Kong epidemic did have antibodies to Hong Kong neuraminidase in their sera and that after vaccination the levels of antineuraminidase did not decrease more rapidly than antibodies to the haemagglutinin. This table: h Kilbourne, E. D., Christenson, W. N. & Sande, M..(1968) J. Virol., 2, 761-762. tS hulman, 3. L., Khakpour, M. & Kilbourne, E. D- (1968) J. Virol., 2, 778-786. 2393A 483- DISCUSSION PAPERS-SESSION II TABLE 1 HAEMAGGLUTINATION-INHIBITION AND NEURAMINIDASE-INHIBITION ANTIBODY LEVELS IN CHILDREN AFTER VACCINATION WITH A2/Aichi/68 VACCINE Number Time after vaccination Prebleed a 4 weeks 30 weeks Haemagglutination-inhibition titres (per ml) 20 80 b 6 900 2 000 Neuraminidase-inhibition titres (per ml) 20 370 c 2 600 770 a Children (2-15 years, median 5 years) were bled before the onset of the Hong Kong influenza epidemic and received 400 CCA units of Eli Lilly Zonomune monovalent vaccine by the subcutaneous route. b Reciprocal of serum dilution giving haemagglutination inhibition. c Reciprocal of serum dilution giving 50% inhibition of 1 unit of neuraminidase activity (as defined by Webster, R. G. & Pereira, H. G. (1968) J. gen. Virol. 3, 201). TABLE 2 FAILURE TO DETECT ANTINEURAMINIDASE ANTIBODIES TO A2 INFLUENZA VIRUSES IN HUMAN SERA COLLECTED BEFORE 1957 a Number of sera with Number of sera with Age in Number positive haemagglutination inhibition positive neuraminidase inhibition 1957 tested (year__s) A2/Japan/305/57 A2VAichi/68 A/Equi-2/63 A2/Japan/305/57 A2/RI/5s A2/Aichi/68 62-91 50 50 50 5 1 1 3-60 50 0 0 0 1 1 1 a These sera were kindly provided by Dr N. Masurel, Leiden, Netherlands. gives no answer to the third question, or to the main problem posed, but shows that more information is required. Another question that arises from the study of the 2 antigens of the Hong Kong influenza virus is whether or not the influenza virus that caused infection in man in the late nineteenth century possessed a neuraminidase antigen similar to that in the present Hong Kong virus. Antisera kindly provided by Dr N. Masurel that were collected before 1957 were examined for evidence of antibodies to A2/Hong Kong neuramini- dase. Only 1 serum gave any evidence of neuramini- dase antibody (Table 2) (then only at a dilution of 1/5) whereas all of the samples from the elderly peoplehadantibodies to A2 influenzahaemagglutinin. From this we could conclude that the neuraminidase of the Hong Kong-like virus present in the late nineteenth century was distinct from that of the present A2/Hong Kong virus but we do not know (1) if neuraminidase antibodies can be detected a long time after initial infection, or (2) if infection or vaccination with an antigenically related neur- aminidase will stimulate antibodies against the neuraminidase of the influenza virus first experienced in childhood as is the case with the haemagglutinin antigen (original antigenic sin). Serum samples from young adults (kindly provided by Dr W. M. Marine) who experienced Al/FM/l/47 as their original influenza virus, were examined for the presence of neuraminidase antibodies to the virus. Of 55 samples tested, 36 showed neuraminidase antibodies (see Table 3). The vaccine record of these patients is unknown. However, they did have measurable levels of antineuraminidase, and by inference we might have expected more positive samples in the pre-1957 sera, for this group of sera from elderly people was selected on the grounds that 484 ANTIGENIC VARIATION IN INFLUENZA VIRUSES 485 TABLE 3 HAEMAGGLUTININ-INHIBITING AND NEURAMINIDASE- INHIBITING ANTIBODIES TO A1/FM/I/47 INFLUENZA VIRUS IN YOUNG ADULTS a Mean titre Test No. positive/ of positiveno. tested samples b (per ml) Neuraminidase-inhibition 36/55 50 Haemagglutination-inhibition 51/55 930 a These sera were kindly provided by Dr W. M. Marine, Atlanta, Ga., USA. b Titres expressed as in Table 1. they possessed A2 haemagglutinin antibodies. The evidence suggests that the virus that occurred in the late nineteenth century possessed a different neura- minidase from that present in Hong Kong influenza virus. I have not tried to provide answers to the 3 points that I have raised-(1) the origin of human influenza strains with one or the other antigen related to animal or avian influenzas, (2) the role of the neuraminidase antigen in protection, and (3) the nature of the neuraminidase antigen of viruses that occurred in humans before 1933. All I have done, I hope, is to provide enough information to stimulate further investigations in these fields. Relationships Between Animal and Human Influenza by M. M. KAPLAN a Prior to the 1957 pandemic only a few workers, led by Shope, paid any attention to animal influenza and its possible relationship to human influenza. The recognition about 15 years ago that fowl plague and equine influenza belonged to the same group of viruses spurred some interest in this field. In July 1957, soon after the onset of the new A2 epidemic along with the vague report from China that epizootics in pigs had also occurred, the World Health Organization sought the opinions of experts throughout the world, including Sir Christopher Andrewes and Sir MacFarlane Burnet, concerning the intriguing possibility that the strain may have emerged from an animal reservoir. Since that time the World Health Organization has been trying to stimulate and co-ordinate studies on animal influenza in different parts of the world: periodic information circulars on animal influenza are sent to interested workers throughout the world. Advances in knowledge in this field during the past few years have been very great. But the following fundamental questions remain unanswered. Are lower animals of any importance as a prime source of the major antigenic shifts in the influenza A group that cause the recurring epidemics and pandemics? If they are, is it because of the emergence of an a Chief, Veterinary Public Health, World Health Organi- zation, Geneva, Switzerland. animal strain, which because of a slight change, is able to spread in man, or do recombinations occur in nature between animal and human strains with the subsequent emergence of a new strain? Do both sorts of events occur? The study of these possibilities is clearly most fascinating and productive, as revealed by several of the preceding papers. Several of the earlier papers have referred to the haemagglutinin (HA) and neuraminidase (E) rela- tionships, and their variability, between the Hong Kong strain and the earlier A2 strains (and even AO and Al) on the one hand and some avian and equine strains on the other. The development during the past few years of precise techniques for studying the neuraminidase enzyme (E) has added a valuable new tool for clarifying this problem. However, the epidemiological significance of these findings remains to be ascertained. Fenner b noted that the appearance of A2 with clear shifts in both HA and E gives considerable support to the idea of emergence from an animal reservoir, rather than representing a shift from Al. It would be interesting to know whether analyses of sera from the aged people investigated included anti-neuraminidase studies, as well as studies of HI. This would give a clearer indication of the relation- b Fenner F. (1968) The biology of animal viruses, New York and London, Academic Press, vol. 2, p. 778. 2393B