An epidemic of acute respiratory tract infection

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Part of Acta Medica Philippina

Title: An epidemic of acute respiratory tract infection
Creator: Chan, Veronica F.; Espiritu-Campos, Lourdes
Language: English
Source: Volume XV (4) April-June 1959
Year: 1958
Subject: Respiratory tract infections; Medicine -- Periodicals
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Fulltext: AN EPIDEMIC OF ACUTE RESPIRATORY TRACT INFECTION AMONG CHILDREN CAUSED BY HA VIRUS: I. SEROLOGY• VERONICA F. CHAN, B.S.HYG., C.P.H . ..• LOURDES ESPIRITU-CAMPOS, M.D., M.P.H. Department of Medical MicrobWlogy, Institute of Hygiene University of the Philippines In late July through mid-September, 1957, the Children's Memorial Hospital (CMH) in Banawe, Quezon City, recorded a total admission of 142 infants and young children with acute respiratory illness given the diagnosis of acute bronchiolitis. The iUness was associated with sudden onset of high fever, restlessness and even sleeplessness. Among younger patients, the manifestation of respiratory distress was observed to bC' more severe while the febrile reaction was slight. On the other hand, the older patients experienced more severe febrile reaction and less respiratory distress. The frequency of cases was highest in the one-month to one year age group (1). ThP CMH and the Virus Laboratory, Depa1tment of Medical Microbiology of the Institute of Hygiene, embarked on a collaborativ(' effort to study the illness. The plan was to study the diseasP simultaneously from the clinical as well as the laboratory points of view. A paper on the clinical aspects of the riisease ha..<; already been published ( 1). Workers abroad have associated a number of rccognizerl viruses as responsible for respiratory illnesses among humans. Aside from the various influenza viruses, agents that have already been cited in reports are the adenoidal-pharyngeal-conjunctival (APC) group of ,viruses (2, 3, 4), hemadsorption <HA) viruses (5), and with less certainty, the crouP-as:sociated (CA) virus (6), respiratory syncytial (RS) virus (7,8), •Th.11 RUlb wa1 aided by u in-ant hum the Nnturnl ScienC'O Keoenreb ··~nt"• .,r th~ llnlvenlty nl the l'hlllpnlneo. lte"'8reh altotmenr No. 1r.. YN•• 1~~~-1%~. 199 2-00 ACTA MBDICA PIDLIPPINA Johns llopkins (Jll) virus (9), and 2060 virus (10). With the ••backdoor approach" (2) as a guide, work has been done employing some of the viruses mentioned. The availability of seed virus in our laboratory dictated the use of the viruses in our investigation. The purpose of this paper is to present a preliminary report of the immunological responses of the cases studied. Isolation of virus will form the basis of succeeding reports. MATERIALS AND METllODS Serum collection: Blood specimens were extracted aseptically, early in illness and approximately two weeks later, from acute bronchiolitis patients at CMH. In both instances, blood samples were iced soon after extraction and while in transit to the virus laboratory. After allowing the blood clot to retract at 4°C., the sera were separated and stored in a -20°C. freezer. Out of the 29 patients ill with acute bronchiolitis that were bled during the acute phase of illness, onJy 17 were subsequently extracted for the convalescent sample. Hence, these 17 paired sera comprised our study group. Comp/£m<nt fU;ation (CF) test: (a) Preparation of antigen: Lyophilized adenovirus tYJ>e 4 was kindly sent to us by Dr. Garrison Rapmund of the Walter Reed Army Medical Research Center, Washington, D.C., U.S.A. In our laboratory, the adenovirus type 4 was grown in bottles of HeLa cells. When the cells showed sufficient degree of cytopathic changes, the bottles were frozen and thawed alternately, and this procedure was performed twice. Fluids were pooled and centrifuged in a refrigerated International Centrifuge ·(Model PR-2) at 2,500 rpm for 20 minutes. The clear supernate was ampouled, sealed and shell frozen and stored at -60°C. This preparation served as CF antigen. Uninfected bottles of HeLa cetls were similarly treated and served as control. (b) Immune serum: Rabbits used for the preparation of immune serum were pre-bled. A series of ten biweekly intravenous injections were scheduled and ten days after the last, SEROLOGY IN ACUTE BRONCHIOLITIS 201 the rabbits were bled by cardiac puncture. Serum was stored at -20°C. (c) Method: Briefly, the method employed 2 units of antigen, 2 full units of complement, 2 units of hemolysin and overnight fixation at 4°C. (11, 12). The highest dilution of serum which exhibited 75% or greater fixation of complement was considered the endpoint. Antibody rise in titer of fourfold or greater was considered significant. Hemaggluti11,Q,tion Inhibition (HI) Test: (a) Preparation of antigens: HA viruses type I and II were sent to us through the courtesy of Dr. R.M. Chanock of the National Institutes of Health, Bethesda, Maryland, U.S.A. Mon.key kidney cells grown in blake bottles prepared at our laboratory according to the technique of Younger et al. (13) were used to propagate the viruses. The monkey kidney tissu~ culture fluids simi1arly treated as that of the adenovirus type 4 were used as antigens. Uninoculated monkey kidney cells prepared in like manner served as controls. Infected chorioallantoic fluids of 12-day old embryonated eggs were sources of antigens for mumps virus and the influenza viruses: A'/FMl/47, A/PHIL/57, A/PR8/34, A/Swine-16 1976/31, A/Denver/57, B/Lee/40 and D/Sendai/52. (b) Immune serum: Immune sera were prepared in big roosters that were pre-bled. Ten days after a series of intravenous injections, the roosters were test-bled. If satisfactory titers were obtained, the roosters were exsanguinated by cardiac puncture. Sera were stored frozen at -2-0°C. without preservative. ·(c) Method: The sera were pre-treated with trypsin in phosphate buffer pH 8.2 and inactivated at 56°C. for 30 minutes (14). Guinea pig erythrocytes were used in the HI test of the HA viruses allowing the erythrocytes to sediment at 4°C. and the test read by the pattern method after 2 hours (5). With the influenza viruses, fowl erythrocytes were employed, sedimentation taking place at room temperature and the test read by the pattern method after one hour. As in the CF test, a fourfoJd or greater rise in antibody titer was considered signific&nt. 202 AcrA. llEDICA PmLIPPINA RESULTS Complement Fim.tion Paired sera obtained from 17 children with acute bronchiolitis were tested for development of complement fixing antibodies against adenovirus type 4. Results are sununarized in Table 1. Only one out of 17 cases showed a significant rise in CF antibody titer. Hemagglutination Inhibition: For the purpose of economy on serum samples, only the convalescent phase of the 17 paired sera were tested by HI test against the following influenza viruses: A'/FMl/47, A/PR8/34, A/Swine-16 1976/Sl, A/Denver/67, and B/Lee/40. In no single instance was an antibody titer greater than 1 :8 of serum diJution. A/Phil/57, an Asian influenza strain isolated in our laboratory and found to be closely related to A/Jap 305/57 was used in HI test with the 17 paired sera. Table 2 presents the data of HI tests performed with the 17 paired sera. In all instances, appreciable antibody titers were shown to exist starting at a serum di1ution of 1 :16. Out of the 17 pairs tested, 1 case showed significant increase in HI antibody titer. HI antibody levels against HA-I (para-influenza 3) and HA-II (para-influenza 1) ·(15) were determined from the 17 cases of acute bronchiolitis. Because of the relation that exists between the HA viruses and Sendai and or mumps virus ( 5). the latter viruses have also been included in the tests. Table 2 summarizes the data. As may be seen from Table 2, out of' 17 cases. 3 showed a significant rise in HI antibody titer against HA-I, and another against HA-II. Of interest to note is the presence of a significant rise in HI antibody titer against A/Phil/57 and HA-II in the same patient ·(No. 10 C.R.). Very low antibody leve1s have been exhibited in all cases against mumps and Sendai viruses. SEROLOGY IN ACUTE BRONCHIOLITIS 203 DISCUSSION With the adenoviruses sharing extensively complement fixing antigens (16) the CF test employing only one type of adenovirus as an antigen proves a serologic test of value in the detection of the existence of an infection caused by any of the other lmown types (2). Studies made by Heubner et al. (2) showed that more than 70% of persons infected with any given strain showed fourfold heterotypic rises against other antigenic types. The likelihood that a type of an adenovirus is the probable etiologic agent of this epidemic of acute bronchiolitis is not great since only one out of 17 cases manifested a significant antibody rise. It must be mentioned that cases of acute bronchiolitis aPpeared at a time when the new A/ Asian influenza/57 strain was circulating. One patient (No. 10) with significant antibody rise against the Asian strain may very well have been a true case of influenza. The HI antibody titers of all cases studied reflect recent exposure to the Asian strain of influenza epidemic of 1957 which occurred before the outbreak of the acute bronchiolitis epidemic. Five out of 17 cases showed a significant rise in antibody titer against the HA viruses, one of the 5 showing a significant rise to both Asian strain and HA-II and is probably a case of double infection. Chanock et al. (5) in a recent publication, incriminated the HA viruses, with HA-I significantly more prevalent among infants and young children with respiratory illnesses such as febrile pharyngitis, acute bronchiolitis and pneumonias, inasmuch as half of the cases studied yielded HA-I virus in their throat swabs. In reexamining the paired sera of cases obtained, we found that not all were ideal specimens for testing as far as tiffie of collection is concerned. In )!Orne of the cases, there is tendency to manifest a significant antibody rise but this probably did not become evident because the acute phase sera were extracted too late in the course of the disease. Because of this limitation, the authors with calculated optimism entertain the possible relation of the HA-I virus as t>tiologic agent in this epidemic of acute bronchiolitis. 204 ACf.A MEDICA PBILIPPINA Further and more extensive controlled studies to corroborate our findings regarding the HA viruses will be the subject of. a future work. SUMMARY A rough immunological screening was performed on 17 acute bronchiolitis cases. Tests employed were CF against adenovirus type 4 and HI against the various influenza viruses: viz., A'/FMl/47, A/ PRB/34, A/Swine-16 1976/31, A/Phil./67, B/Lee/40 and D/ Sendai/52; the mumps virus, HA-I and HA-II. , The likelihood that a type of an adenovirus as the probable viral etiologic agent of this epidemic of acute bronchiolitis is not great since only one of 17 cases gave a significant antibody rise in titer. The possible role of the various influenza viruses have been eliminated. The possible etiologic relation between the HA-I virus and acute bronchioJitis is suspected based on the observation that four (4) of the seventeen (17) cases tested or about one-fourth showed positive results, but further and extensive controlled study is needed. ACKNOWLEDGMENTS Grateful acknowledgment is due to Dr. Fe del Mundo, Director, Children's Memorial Hospital, who made possible the collection of clinical specimens. The authors wish to thank Dr. Potenciano R. Aragon, Head, Department of Medical Microbiology, Institute of Hygiene, for his suggestions in writing this paper. The technical assistance of Mrs. Gorgonia T. Ocampo is greatly appreciated. SEROLOGY IN ACUTE BRONCHIOLITIS 205' REFERENCES 1. MUNDO, F. de!, SORIANO, L., AFRICA-JULIANO, L., MATAWA· RAN, A., and SIAZON P.: An Epidemic of Bronchiolitis in Infants in 1957, Jour. Phil. Med. Ass., 12:729-735, Dec. 1958. 2. HEUBNER, R.J., ROWE, W.P., WARD, T.G., PARROTT, R.H., and BELL, J.A.: Adenoidal-Pharyngeal-Conjunctival Agents; a Newly Recognized Group of Common Respiratory System Viruses, Nev.England Jour. Med., 251:1077-1086, 1954. 8. PARROTT, R.H., ROWE, W.P., DEUBNER, R.J., BERNTON, H.W., and McCULLOUGH, N.B.: Outbreak of Febrile Pharyngitis and Conjunctivitis Associated with Type 3 APC Virus Infection, New England Jour. Med., 251:1087-1090, 1954. 4. HILLEMAN, M.R., and WERNER, J.B.: Recovery of New Agent from Patients with Acute Respiratory Illness, Pro. Soc. Exper. Biol, and Med., 85:183-188, 1954. 6. CHANOCK, R.M., PARROTT, R.H., COOK, K., ANDREWS, B.E., BELL, J.A., REICHELDERFER, T., K.APIKJAN, A.Z., MASTROTA, F.M., and HEUBNER, R.J.: Newly Recognized Myxoviruses from Children with Respiratory Diseases, The New England Jour. Med., 6:207-213, Jan. 30, 1958. 6. CHANOCK, R.M.: Association of a New Type of Cytopathogenic Myxovirus with Infantile Group, The Jour. Exper. Med., 4:555575, Oct. 11, 1956. 7. CHANOCK, R.M., REIZMAN, B., and MYERS, R.: Recovery from Infants with Respiratory lllness of a Virus related to Chimpanzee coryza Agent (CCA) I. Isolation Properties and Characterization, Am. Jour. Hyg., 63:281-290, Nov. 1957. 8. CHANOCK, R.M., and FINBERG, L.: Recovery from Infants with Respiratory Illness of a Virus related to Chimpanzee Coryza Agent (CCA). II. Epidemiologic aspects of Infection in Infants and Young Children, Am. Jour. Hyg., 63:291-300, Nov. 1957. 9. PRICE, W.H.: The Isolation of a New Virus Associated with Respiratory Clinical Disease in Humans, Proc. Nat. Acad. Sci. 42:892896, 1956. 10. PELON, W. MOGABGAB, W. J., PHILIPS, I.A., alid PIERCE, W.E.: A Cytopathogenic Agent Isolated From Naval Recruits with Mild Respiratory Illnesses, Proc. Soc. Exper. Biol. and Med., 94:262~267, 1957. 111.: BENGTSON, I.A.: Complement Fixation in the Rfokettsial Diseases Technique of the Test, Pub. Health Rep., 59:402-406, 1944. ACTA -MEDICA PBILIPPINA 12. BEEMAN, E.A., HEUBNER, R.S., and COLE, R.M.: Studies of Coxsackie Viruses. Laboratory Aspects of Group A Viruses, Am. Jour. Byg., 66:83-107, 1962. 13. YOUNGER, J.S.: Monolayer Tissue Cultures. I. Preparation and Standardization of Suspensions of Trypsin-Dispersed Monkey Kidney Cells, Proc. Soc. Exper. Biol. and Med., 86:202-206, 1964. 14. KEITH, E.J.: "Influenza", American Public Health Diagnostic Pro· ceduree for Viruses and Rickett8ial Diaeases, 2nd ed. New York, 1966. 10. ANDREWES, C.H., BANG, F.B., CHANOCK, R.M. and ZHDANOV, V.M.: Para-Influenza Viruses 1, 2, and 3: Suggested Names for Recently Described Myxoviruses, Virology, 8:129-130, 1969. 16. ROWE, W.P., HEUBNER, R.J., HARTLEY, J.W., WARD, T.G. and PARROTT, R.H.: Studies on the Adenoidal-Pharyngeal-Conjunctlval (APC) Group of Viruses. Am. Jour. Byg., 61:197-218, March 1965. SEROLOGY IN ACUTE BRONCBIOLITIS 207 TABLE 1 RESULTS OF CF TESTS WITH 17 BRONCHIOLITIS CASES TO ADENOVIRUS TYPE 4 1. J. c. (Male) 2. G. C. (Male) 3. H. C. (Female) 4. c. c. (Female) .'i. J. c. (Male) 6. L. H. (Female) 7. A. J. (Male) 8. M.L. Jr. (Male) 9. L. R. (Female) w. C.R. (Female) 11. J. s. (Male) 12. E. SJ. (Female) 13. L. SJ. (Female) 14. B. S. (Male) 15. JM. S. (Male) 16. BB. V. (Male) 9.4.57 23 8-25-57 9. 1-57 11 9- 9.57 11 8-20-57 12 8-27-67 II 9- 1-57 8-27-67 12 8-31-67 8-24-67 9- 1-57 8-30-57 11 8-28-67 8-25-57 8-21-57 "-·~?~"~,:;~~)~~- _ _l_~--25-~7. J_ 0 - Less than 1 :8 dilution. • 20 • 15 8 13 18 18 6 11 8 13 3 10 10 19 8 15 • 11 • 15 • 18 • ..!_2_ 1:266 1:256 0 1:64 0 1:8 1:16 1:8 0 __o_ x -The patient was first admitted at the CMH June 15, 1957, and re~~em~~~!n~i~asthbie~i8'roer l~'rt11~~dtss~!~~~b{in 4Se~~~b:~. is~i~'9stt: second blood sample was withdrawn. 208 . AC?A HIIDICA PBILIPPINA TABLE 2 m ANTIBODY TITERS TO INFLUENZA ASIAN STRAIN A/PHIL/67, HA VIRUS TYPE I AND BA VIRUS TYPE II Patient No. A/PhiV67 HA-I HA-ll 1:286 1:8 1:266 1:16 1:18 1:82 1:18 1:16 1:82 1:16 1:82 1:64 1:82 1:32 1:64 1:16 1:266 0 1:128 1:32 1:64 1:128 1:128 1:612 1:82 0 1:32 0 1:266 1:64 1:32 1:128 1:266 1:82 1:128 1:128 1:266 1:128 1:16 1:8 1:16 1:16 1:8 1:16 10 0 • 0 1:82 0 1:64 11 1:266 1:64 1:82 1:128 1:82 1:32 12 1:64 1:128 1:32 1:64 1:128 1:64 18 1:64 1:8 1:16 1:32 1:8 1:16 " 1:32 1:8 1:8 1:32 1:128 0 .. 1:64 1:8 1:8 1:64 1:8 1:8 •• 1:64 1:128 1:32 1:64 1:128 1:32 17 1:64 1:32 1:16 1:64 1:16 1:16 0-Leea than 1:8 dilution.
pages: 199-208