Respiratory syncytial virus or influenza?

THE LANCET I

I

Respiratory syncytial virus or influenza? D M Fleming, K W Cross Summary We compared data from clinical surveillance of acute respiratory infections in elderly people reported by a network of sentinel general practitioners, virus isolate data for respiratory syncytial virus (RSV) and influenza A and B viruses, and registered deaths, during the past four winters. Cases of acute respiratory disease (ARD) and deaths (3-week moving averages) showed strong similarity in timing. Periods of high incidence of ARD and numbers of deaths were followed within 2-3 weeks by reports of high numbers of RSV or influenza virus isolates. The epidemic period in December, 1989, was followed by high numbers of both virus isolates, but in all other epidemic periods one virus predominated. The lethality of epidemics due to RSV or influenza A or B varied both within and between years. These analyses suggest that RSV is as important as influenza viruses in causing morbidity and excess deaths among elderly people. RSV was prevalent at the end of each of the past four years in the UK; its importance as a cause of excess deaths would therefore be concealed in any analysis based on several years' experience. By contrast, the inconsistency of epidemics due to influenza viruses makes excess deaths due to these viruses easily recognisable. Integrated clinical and viriological surveillance is essential if epidemics due to RSV, influenza virus, or any other virus are to be correctly interpreted.

Lancet 1993; 3 4 2 : 1 5 0 7 - 1 0 Introduction T h e influence o f season o n m o r t a l i t y has b e e n r e c o g n i s e d since the t i m e o f Hippocrates.1 M o s t o f t h e studies i n c l u d e d in A l d e r s o n ' s 1988 r e v i e w 2 h a d c o n c e n t r a t e d o n d e a t h s f r o m cardiovascular disease a n d o n t e m p e r a t u r e a n d climatic variables; a few also took a c c o u n t o f r e s p i r a t o r y m o r b i d i t y . T h o u g h reliable data were available for all d e a t h s a n d for t e m p e r a t u r e , t h e r e were virtually n o data o n a c u t e r e s p i r a t o r y illness. T h u s , proxies for m o r b i d i t y were used, i n c l u d i n g r e s p i r a t o r y deaths, h o s p i t a l a d m i s s i o n s for p n e u m o n i a , a n d sickness a b s e n c e certificates. A s t u d y o f m o r t a l i t y in f o u r c o u n t r i e s a s h o w e d t h a t f l u c t u a t i o n s in t h e incidence a n d severity o f r e s p i r a t o r y infections c o u l d affect m o r t a l i t y f r o m n o n - r e s p i r a t o r y as well as r e s p i r a t o r y causes. C u r w e n et al 4 r e p o r t e d t h a t d e a t h s f r o m all causes increased at t h e s a m e t i m e as clinical r e p o r t s o f i n f l u e n z a like illness in t h e weekly r e t u r n s service o f t h e Royal College of G e n e r a l P r a c t i t i o n e r s ( R C G P ) . W e have s h o w n , u s i n g data f r o m these s e n t i n e l practices in E n g l a n d a n d W a l e s , t h a t t h e r e is a relation, possibly causal, b e t w e e n total acute r e s p i r a t o r y illness in t h e elderly a n d m o r t a l i t y : W e p r e s e n t h e r e weekly data for t h e past f o u r w i n t e r s o n acute r e s p i r a t o r y diseases in t h e elderly, total d e a t h s , a n d

Birmingham Research Unlt/RCGP, LordswoodHouse,54 Lordswood Road, Harbome, Birmingham B17 9DB, UK (D M FlemingFRCGP, K W Cross PhD) Correspondenceto: Dr D M Fleming

Vo1342 • December 18/25, 1993

isolations o f r e s p i r a t o r y syncytial v i r u s ( R S V ) a n d i n f l u e n z a viruses. W e were n o t c o n c e r n e d w i t h elderly p e o p l e living i n i n s t i t u t i o n s , for w h o m t h e p a t h o g e n i c i t y o f R S V a n d of i n f l u e n z a is widely r e c o g n i s e d . 6~

Methods Weekly data for the past 4 winters (between week 47 of one year and week 18 oft.he next; total 24 weeks) were collected for all r e g i s t e r e d deaths (at age 1 year or older), numbers of isolates of influenza A, influenza B, and RSV, and rates of new episodes of all a c u t e respiratory disorders in elderly people (65 years and older) as reported to general practitioners. (Since 1992 was a 53-week year, the 24-week study period for 1992/93 was week 47, 1992, to week 17, 1993). Total deaths (all ages and all causes) were used for this study; 81% of all deaths are of people of 65 and older. T h e effect on deaths from influenzal illness extends beyond deaths attributed to respiratory disease: ,s Virus isolate data were provided by the Communicable D i s e a s e Surveillance Centre (CDSC). These data are presented according to the time the report of a positive isolate is received at the CDSC rather than the time the sample was taken; this practice leads to a delay of about 3 weeks. 9 Respiratory illness data were taken from the weekly returns service of the R C G P TM and were based on an average w e e k l y population of about 500 000, 15 % of whom were aged 65 or older. About half of new cases of acute respiratory disease (ARD) in the elderly are attributed to acute bronchitis during the winter. ARD rates as used here refer to patients consulting general practitioners; the decision to consult the general practitioner is an indication of the clinical severity. T h e data sets were examined graphically for factors that might explain differences in the pattern of winter morbidity and mortality from one year to another. For each winter, 3-week moving a v e r a g e s of the total rates for new episodes of ARD and of the numbers of deaths (all ages) were plotted by week. We set arbitrary n u m b e r s for virus isolates; periods with rates above a 3-week moving a v e r a g e of 500 for RSV and 100 for influenza A and B combined w e r e defined as peak periods.

Results T h e n u m b e r s o f R S V isolates r e p o r t e d to t h e C D S C were 7857 in t h e w i n t e r o f 1989/90, 6950 for 1990/91, 10 382 f o r 1991/92, a n d 9866 for 1992/93. T h e weekly p a t t e r n p r e s e n t e d as g r a p h s o f 3-week m o v i n g a v e r a g e s is s h o w n i n figure 1. T h e r e w e r e d i f f e r e n c e s year b y year i n t h e rate o f i n c r e a s e to a n d d e c l i n e f r o m t h e peak, a n d also in t h e d u r a t i o n o f peak p e r i o d s ( 3 - w e e k m o v i n g a v e r a g e > 500 isolates). H o w e v e r , t h e peak o c c u r r e d at a l m o s t t h e same t i m e each year; an i n c r e a s e was d e t e c t a b l e b y week 49. R e p o r t e d isolates o f i n f l u e n z a A a n d B totalled 2914 i n 1989/90, 1854 in 1990/91, 1684 in 1991/92, a n d 837 in 1992/93. I n 1989/90, a l m o s t all t h e isolates w e r e i n f l u e n z a A a n d in 1990/91 a l m o s t all were i n f l u e n z a B. I n t h e o t h e r t w o w i n t e r s t h e r e were a p p r o x i m a t e l y e q u a l n u m b e r s o f t h e t w o v i r u s e s . F i g u r e 1 s h o w s 3 - w e e k m o v i n g averages o f i n f l u e n z a A a n d B v i r u s isolates c o m b i n e d . T h e r e were d i f f e r e n c e s year b y y e a r in t h e t i m i n g o f t h e h i g h e s t n u m b e r o f isolates a n d the d u r a t i o n o f p e a k p e r i o d s (3-week m o v i n g a v e r a g e > 100 isolates). R a t e s of A R D a n d t h e n u m b e r o f d e a t h s r e g i s t e r e d i n e a c h week are also p r e s e n t e d as 3 - w e e k m o v i n g averages for

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THE LANCET

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each of the four winters (figure 2). In 1989/90, the highest incidence of A R D occurred in week 49 and the highest number of deaths in week 51; the numbers of virus isolates for both R S V and influenza virus increased rapidly from week 52 and both reached a maximum in week 3. In 1990/91, both A R D and deaths showed two peaks; the A R D peak in week 1 was substantially higher than that in week 7, whereas the two peaks in deaths were similar. T h e maximum number o f RSV isolates was reported in week 3 (peak period 2-4) and influenza isolates in week 12 (6-14). T h e r e were again two peaks of deaths and o f A R D in 1991/92. T h e higher peaks of both occurred in week 2, at the same time as highest number of RSV isolates (figure 1). A small secondary peak of A R D and deaths at week 6 was followed by the maximum number of influenza isolates in week 9, though the 3-week moving average was above 100 isolates from week 3 onwards. A R D rates were slightly higher in this winter and in 1992/93 than in the previous two winters because of the transfer from paper to computerbased recording systems for the Fourth M o r b i d i t y Survey in General Practice. In 1992/93, the substantial peaks of A R D (week 1) and deaths (week 2) were followed by the maximum n u m b e r of RSV isolates in week 7 (peak period 2-10). D u r i n g the secondary rise (weeks 8-16), the incidence of A R D and deaths maintained a close relation, though the increases in both were seen later than in previous years, occurred over a longer period, and showed lower m a x i m u m values. Influenza isolates "were greatest in week 16 (peak period 14-18). During the first 5 weeks of the year, there were on average only 12 influenza isolates per week.

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Vol 342 • D e c e m b e r 18]25, 1993

rI'HELANC~'I"

Discussion This study compared published statistics that are independent of each other. The death data are presented as 3-week moving averages to smooth out irregularities in death registration due to bad weather and to the Christmas holiday period. Data from all causes were used since influenza deaths are often hidden among other causes. 4,1t The virological data are collected for routine clinical purposes and are largely based on samples from children. In 1989/90, when there was an influenza epidemic, about 63% of positive influenza isolates were from children and only 13 % were from people aged 65 and older; the corresponding proportions for RSV were 98% and 1%. During that period there were more than twice as many isolates of RSV as of influenza. In subsequent years there were even greater differences, for example, a tenfold difference in 1992/93. The clinical signs of acute respiratory illness provide the basis for diagnosis of ARD but they are not related to specific pathogens; it may be impossible to distinguish clinically between influenza and RSV infections in elderly people. ~-a I n all four winters there were substantial numbers of RSV isolates and in three, the threshold value of S00 isolates was reached in week 1 of the year. In 1991/92, the increase was evident a little earlier and the m a g n i t u d e of the peak was greater. This pattern was not seen for influenza isolates; the data showed increased influenza activity in weeks 51 to 7, 6 to 14, 3 to 12, and 14 to 18, which suggests random occurrence during winter. In all four years, there were recognisable epidemics of ARD and consistently matching concurrent periods of increased deaths. Every peak observed was followed immediately by peaks in isolates of either RSV or influenza viruses~ with the exception of late 1989, when RSV and influenza peaked together. T h e peak period for morbidity and mortality in early January every year cannot be explained by influenza. In many laboratories, samples submitted for culture would have been tested for both RSV and influenza viruses. I f influenza viruses were prevalent, it is unlikely they would have been missed. T h e likelihood of death in epidemics due to RSV or influenza A or B virus varied substantially within and between years. I n 1989/90, the epidemic of respiratory illness and numbers of deaths were linked to both viruses. In 1990/91, the apparent RSV epidemic was associated with more morbidity but fewer deaths than the later influenza B epidemic. In 1991/92, the increase in numbers of deaths and incidence of respiratory illness occurred when reports of RSV were maximum, though reports of influenza viruses rose a few weeks later. In 1992/93, there was a greater incidence of ARD and n u m b e r of deaths in the early weeks of 1993, when RSV was prevalent, than when influenza isolates increased later in the winter. It is possible that a combination of particulate matter in the atmosphere and cold (just above 0°C), damp conditions optimises the spread of RSV, which is thought to be spread in large droplets. ~2 T h e countries of mainland Europe experience colder winters than the U K yet report less excess winter mortality. ~s.t, There is a relation between particulate pollutants and respiratory deaths. ~s We draw three main conclusions from this study. First, RSV is an important respiratory pathogen for the elderly in the community. This study cannot distinguish between influenza and RSV as the m6re important cause of epidemics of respiratory illness and of deaths in winter. We Vol 342 • December 18/25, 1993

have concentrated on these two viruses, but it is possible that other pathogens with similar epidemic characteristics are involved, and a search for these is equally important. However, we already know that RSV is a serious respiratory pathogen for young children t6 and for people living in institutions or nursing homes. 6-a A study of the contribution of RSV to epidemics of acute respiratory illness among the elderly in the community is needed. Our second conclusion is that the calculation of excess deaths over an expected n u m b e r derived from previous experience is unsatisfactory. RSV has a reputation for its consistent appearance at the beginning of the year. l~.ts Numbers of deaths are also consistently greatest at this time of year. Thus, any formula for deriving expected deaths that is based on accumulated experience over several years will derive expected numbers very similar to the observed number, and the potential of an epidemic illness to cause excess deaths is concealed. Outbreaks of influenza do not occur at the same time each winter; thus, excess numbers of deaths attributable to influenza are obvious. Thirdly, we conclude that virological surveillance is inadequate. It is recognised that deaths attributable to influenza occur predominantly in the elderly but there is barely any virological investigation of elderly people. Throughout the whole winter of 1992/93, there were only 239 positive influenza isolates for England and Wales, and only 6 of these were from elderly people. We need a system in which there is adequate virological surveillance for populations in which the diseases are monitored. T h e main risk from influenza-like illnesses is in elderly people, most of whom are treated in general practice. Incorrect attribution of epidemics of respiratory disease to influenza, rather than RSV infection, may explain the difficulty of assessing the efficacy of influenza vaccination in clinical studies that have no virological validation. We thank the Office of Population Censuses and Surveys for mortality data and the CDSC for virus isolate data. Morbidity data are available as part of the routine monitoring activities of the Birmingham Research Unit/RCGP. We also thank our network of recording practices; Dr D L Crombie for comments during preparation of this paper; and the Department of Health for support.

Roforon©os 1 Adams F. The genuine works of Hippocrates. Translated from the Greek with a preliminary discourse and annotations. London: Sydenham Society, 1849: 203. 2 Alderson MR. Mortality, morbidity and health statistics. Basingstoke: Macmillan, 1988: 122-25. 3 Anderson M, Le Riche WH. Cold weather and myocardial infarction. Lancet 1970; i: 291-96. 4 Curwen M, Dunnell K, Ashley J. Hidden influenza deaths 1989-90. Population Trends 1990; 61" 31-33. 5 Fleming DM, Cross KW, Crombie DL. Respiratory illness and mortality in England and Wales. Fur J )gpidemiol (in press). 6 Garvie DG, Gray J. Outbreak of respiratory syncytial virus infection in the elderly. B M J 1980; 281: 1253-54. 7 Agius G, Dindinaud RJ, Biggar RJ, et al. An epidemic of respiratory syncytial virus in elderly people: clinical and serological findings. ff &fed Viro11990; 30:117-27. 8 Osterweil D, Norman D. An outbreak of an influenza-like illness in a nursing home..7.4m Geriatr Soc 1990; 38: 659-62. 9 Joseph CA, Dedman D, Fern K, Chakraverty P, Watson JM. Influenza surveillance in England and Wales: November 1991-June 1992. Commun Dis Rev 1992; 2: 149-52. 10 Fleming DM, Norbury CA, Crombie DL. Annual and seasonal variation in the incidence of common diseases. Occasional Paper 53. London: Royal College of General Practitioners, 1991. 11 Office of Population Censuses and Surveys. Mortality statistics by cause 1986 (England and Wales). Series DH2 no 13. London: HM Stationery Office 1988.

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THE LANCET 12 Graman PS, Hall CB. Nosocomialviral respiratory infections. Semin Respir Infect 1989; 4: 253--60. 13 Curwen M. Excess winter mortality: a British phenomenon. Health Trends 1990; 221 169-74. 14 Kunst AE, Looman CWN, MackenbachJP. The decline in winter excessmortality in the Netherlands. Int Epiderniol 1990; 20:971-77. 15 Schwartz J, Dockery DW. Increased mortality in Philadelphia associatedwith daily air pollution concentrations. Am Rev Resplr Dis 1992; 145."600-04.

16 Martin AJ, Gardner PS, McQuillin J. Epidemiologyof respiratory viral infection among paediatric inpatients over a six year period in northeast England. Lancet 1978; i: 1035-38. 17 Noah ND. Cyclicalpatterns and predictability in infection. Epideraiol Infect Dis 1989; 102-"175-90. 18 Dab W, Quenel P, Cohen JM, Hannoun C. A new influenza surveillance system in France (the Ile-de-France "GROG"): 2 validity of indicators (1984-1989). Eurff Epidemio11991; 7: 579-87.

Induction of nitric oxide synthase in asthma Qutayba Hamid, David R Springall, Valentina Riveros-Moreno, Pascal Chanez, Peter Howarth, Anthony Redington, Jean Bousquet, Philippe Godard, Stephen Holgate, Julia M Polak Summary

Nitric oxide (NO) is a mediator of vasodilatation and bronchodilatation synthesised from L-arginine by the enzyme NO synthase, which is either constitutive or induced by lipopolysaccharides and/or cytokines. The presence and function of NO synthase in normal or diseased lung is not yet clear. Asthma is characterised by bronchial hyperresponsiveness, epithelial damage, inflammation, and increased cytokine production. To investigate the presence of NO synthase in asthma, we immunostained bronchial biopsies from nonsteroid-treated people with asthma and non-asthmatic controls with specific polyvalent antisera to purified inducible NO synthase and to a selected peptide sequence of the same enzyme. Immunoreactivity was seen in the epithelium and some inflammatory cells in 22 of 23 biopsies from people with asthma, but in only 2 of 20 controls. To assess the relation of cytokines to NO synthase induction, bronchial epithelial cells in culture were stimulated with tumour necrosis factor (TNF~O. Inducible enzyme immunoreactivity was found only in the treated cells. The existence of inducible NO synthase in human lungs suggests that increased production of NO, probably induced by cytokines, may be relevant to the pathology of asthma.

Lancet 1993; 3 4 2 : 1 5 1 0 - 1 3

introduction N i t r i c o x i d e ( N O ) plays a m a j o r p a r t in cell-to-cell c o m m u n i c a t i o n . 1 I s o f o r m s o f N O synthase, the e n z y m e t h a t generates N O f r o m L - a r g i n i n e , 12 have b e e n f o u n d in e n d o t h e l i u m a n d n e u r o n s , a,4 A n o t h e r , t r a n s c r i p t i o n a l l y i n d u c e d by cytokines, has b e e n identified in m a n y cells i n c l u d i n g h e p a t o c y t e ss a n d m o u s e m a c r o p h a g e s 6 f r o m w h e r e it has b e e n ~loned. 7'8 T h e i n d u c t i o n can b e p r e v e n t e d by glucocorticoids. 9 I n lungs, N O causes b r o n c h o dilatation la and vasodilatation, 11a2 and is d e t e c t a b l e in exhaled air. 13 I t m a y also act as a n e u r o t r a n s m i t t e r o f n o n - a d r e n e r g i c , n o n - c h o l i n e r g i c nerves, t4 T h e i n d u c t i o n of N O s y n t h a s e b y i m m u n o m o d u l a t o r s has b e e n r e p o r t e d in rat lung, is w h e r e it m a y be involved in lung injury, t6 H o w e v e r , t h e r e has b e e n n o r e p o r t o f i n d u c i b l e N O s y n t h a s e in h u m a n lungs. A s t h m a is c h a r a c t e r i s e d b y bronchial infiltration w i t h l y m p h o c y t e s , e o s i n o p h i l s , m a c r o p h a g e s , m a s t cells, a n d m y o f i b r o b l a s t s ; collagen d e p o s i t i o n b e n e a t h t h e b a s e m e n t m e m b r a n e ; a n d epithelial-cell d e s q u a m a t i o n , tT-z° R e c e n t r e p o r t s have s h o w n local p r o d u c t i o n o f cytokines, 2t,z2 w h i c h stimulate i n f l a m m a t o r y a n d b r o n c h i a l epithelial cells to s y n t h e s i s e various m e d i a t o r s . 23 W e i m m u n o s t a i n e d b r o n c h i a l biopsies w i t h specific antisera to d e t e r m i n e w h e t h e r i n d u c i b l e N O s y n t h a s e is p r e s e n t in t h e b r o n c h i a l m u c o s a , a n d also e x a m i n e d t h e e x p r e s s i o n o f N O s y n t h a s e in c u l t u r e s o f n o r m a l b r o n c h i a l epithelial cells b e f o r e a n d after s t i m u l a t i o n w i t h a c y t o k i n e , t u m o u r necrosis factor ( T N F a ) .

Patients

and methods

Antisera

National Heart and Lung Institute, Dovehouse Street, London (Q Harold MD), Department of Hlstochemlstry, Royal Postgraduate Medical School, Du Cane Road, London, UK (D R Springall PhO, ProfJ M Poiak osc); Wellcome Research Laboratories, Beckenham, Kent, UK (V Riveros-Moreno PhO);Service des Maladies Resplratolres, H6pltal Arnaud de VIIleneuve, 34059 Montpelller, France (P ChanezMD,ProfJ BousquetPhD,Prof P GodardMo); Immunopharma¢olo~ Group, Department of Medicine, Southampton General Hospital, Southampton, UK (Prof S Holgate DSc,P HowarthDM,A Redington,MRCP) Correspondence to: Dr David R Springall, Department of Histochemistry, RoyalPostgraduateMedical School, Du Cane Road, LondonW12 ONN,UK

1510

Wistar rats were injected intraperitoneally with endotoxin (Sigma, Poole, UK); 6 hours later, the lungs were removed and N O synthase partly purified as described4 up to the ADP-eluate step. Semi-preparative acrylamide gel electrophoresis was done with the ADP-eluate, the 135 kDa protein band excised, the protein eluted and rabbits imrnunised by subcutaneous injection wth 0.25 m L o f t solution containing gel-purified enzyme (30 ~tg) in Freund's complete adjuvant, with a booster 21 days later (the same amount of antigen in Freund's incomplete adjuvant). Serum was obtained 4 weeks later. The published sequence of macrophage NO synthase' was analysed to predict the possible surface exposure of peptide sequences and therefore their likelihood to be immunogenic.24 Several peptides with low homology to the brain constitutive enzyme were synthesised. The peptide sequence Q N G S P Q L L T G T A Q N V P E S L D K L H V T C chosen for this study corresponds

Vo1342 December 18/25, 1993 •