rCP/CDS/OlO
ENGLISH ONLY
!~FOR ARTHROPOD- AND RODENT-BORNE VIRUSES
~RKrNG
GROUP ON DEVELOPMENT OF VACCINES
Convened by tbe REGIONAL OFFICE FOR THE WESTERN PACIFIC OF THE WORLD HEALTH ORGANIZATION
Sendai. Japan 28-31 August 1984
Not for sale Printed and distributed by the Regional Office for the Western Pacific of the World Health Organization Manila. Philippines February 1985
NOTE The views expressed in this report are those of the Members of the Working Group and do not necessarily reflect the policies of the Organization.
This report has been prepared by the R.,ional Office for the We.tern Pacific of the World Health Organizatioa for aovernaents of Member States in the Re~ion and for those who participated in the Workiag Group on Development of Vaccines for Arthropod- and Rodent-Borne Viruses, which was held in Sendai, Japan, from 28 to 31 August 1984.
CONTEIiTS
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INTRODUCTION .... " "
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2. 3.
HAEMORRHAGIC FEVER WITH RENAL SYNDROME ••••••••••• JAPANESE ENCEPHALITIS ............ " .................. " ................ " .. ..
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DENGUE HAEKORRHAGIC FEVER .••••••••••••..•••.•..•• RECOMMENDATIONS ................ """........................ " .. "".. ".... " ..
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5.1 5.2 5.3
5.4 5.5 5.6
Vaccines ••..•.•••.............•....•••...... Vaccine development by modern biotechnology •• Laborator, dia~nosis ••••••.•••••••••••••.••• Epidemiolo., and surveillance .•..••••••••••. Research on pathophysiology ••••.••••••.••••• Newsletter .................................... " ...... " .................... .. AGENDA ........ "" .. "........ ".. ",. .. ".. ".................. ""
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Annex 1
Annex 2 Annex 3 -
LIST OF MEMBERS, CONSULTANTS, OBSERVERS AND SECRETARIAT "" .......... " ...................... " .. ..
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TECHNICAL BASIS FOR RECOMMENDATION ON HAEMORRHAGIC FEVER WITH RENAL SYNDROME" " ...... " " ......................... .
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1.
INTRODUCTION
The WorkinR Group on Development of Vaccines for Arthropod- and Rodent-Borne Viruses met in Sendai, Japan, from 28 to 31 August 1984. The Group was welcomed by Dr H. Nakajima, ReRional Director, WHO ReRional Office for the Western Pacific, who .tress~ that den~ue haemorrhagic fever and Japanese encephalitis were a~thro~d-bor.e virus diseases of increasing public health concern in both the Western Pacific and South-East Asian reRions. Haemorrha.ic fever with renal syndrome (HFRS) was s serious public health problem in China, the Republic of Korea and among laboratory workers in Japan. The objectives of the meetin~ were to determine ways of strengthenin~ cooperation in the development of effective dengue and Hantaan virus vaccines and of a less costly vaccine for Japanese encephalitis. The terms of reference for the Working Group ware to review and discuss progress in the development of arthropod- and rodent-borne virus diseases and to formulate a plan for the development of (1) monovalent and polyvalent JE vaccines; (2) research on component vaccines of dengue virus; (3) research on aantaan virus vaccines. Dr Koinuma of the Ministry of Health and Welfare, Japan, in welcoming the Group, contrasted the situation or Japanele encephalitis before the Second World War, when the annual incidence approached 5 cases per 100 000 population with a fatality rate of about 30%, with the current situation where the disease was virtual Iv controlled in Japan. The Group elected Dr Akira Ova Chairman, Professor Ho Wang Lee Vice-Chairman and Dr Barry Gorman Rapporteur. The agenda and list of participants are attached as Annexes 1 and 2.
2.
HAEMORRHAGIC FEVER WITH RENAL SYNDROME
Haemorrha~ic fever with renal syndrome (HFRS) is a major public health problem in China, the eastern part of USSR, the Republic of Korea and among laboratory workers in Japan. There have a180 been a number of infections recorded in European countries.
The disease is caused by a number of serologically related viruses. From a recent biochemieal analysis, the viruses contain genomes consistin~ of three se~ents of single stranded RNA and have aD RNA-protein structure consistent with classification in the fa.i1y Bunyaviridae. Numerous viruses have been described but not all have been associated with the disease. Studies using monoclonal antibodies have revealed significant antigenic variation within the group of viruses. Conservation of the 3' terminal sequences of each of the genome segments in all viruses examined has led to the proposal that thev be classified as a distinct ~enus of the family Bunyaviridae.
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II . In the natural history of the viruses, there is a close association w1th rodents and high risks of infection occur in situations where man is
exposed to the excreta of infected rodents. In all countries where studies of an~ibodies in rodents have been conducted, there is evidence of both restricted and widespread infection. In some countries, antibody prevalence rates in urban rats are extremely hi~h. There have been infections recorded in man from laboratory-housed animals indicating that exchan2e of animals or export of laboratory animals may constitute a health risk to laboratory workers. The incidence of the disease varies ~reat1y. In 1982, more than 60 000 cases were recorded in China while in the same year, about 500 cases were recorded in the Republic of Korea. In Japan, the positive antibody rate in selected serum bank collections is less than 1% compared with 4.5% in a sample of sanitation workers, e.2. ~arba~e collectors. The groups at hi2hest risk are those working in environments where the incidence of infected rats is hi 2h. In Japan the hi2hest infection rates were found amon~ people involved in the analysis of soil samples, where exposure to excreta in dust may be frequent throu~h sampling and analysis of dumped garba~e and soil. No vaccines are available to combat HFRS and strategies for the development of effective vaccines are complicated by the lack of a suitable animal model system, by the anti~enic variation among the viruses suspected of causing the disease, and potentially complicated by the fact that the viruses contain se2mented genomes. To avoid the possibilities of genetic reassortment between natural viruses and a live vaccine, it will probab1v be necessary to develop inactivated or sub-unit vaccines (see Annex 3). The immunofluorescence antibody staining test (IFA) has found routine use in diagnosis of infection with Hantaan and related viruses. The test is group-reactive but its efficiency appears to be related to the antigen used in the test. ELISA tests have been introduced, which are more sensitive than IFA. For diagnostic purposes, IAHA (immune adherence haemaRRlutionation) and CF (complement fixation) tests have been applied and show promise of being added to the battery of serological procedures (see Annex 3). The application of panels of monoclonal antibodies to Hantaan-re1ated viruses has sug2ested clusters of antigenic complexes within the group. One application of plaque reduction neutralization tests has divided the viruses into four antigenic sub-complexes.
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3.
JAPANESE ENCEPHALITIS
The changin~ epidemiological pattern of Japanese encepha1iti. (JE) was described in the report of the Working Group on the Prevention anrl Control of Japanese Encephalitis, held in Tokyo, Japan, on 19-21 December 1983. The JE virus is a major cause of epidemic encephalitis and presents significant problems in the People's Republic of China, Thailand, Nepal, India and in recent years in the Republic of Korea. In Japan, Burma and Sri Lanka, there are fewer than 100 cases per year.
3
It is perhaps too soon to an~icipale that the recommendations of the previous Workin~ Group in re~ard to staQdard~ation of inactivated vaccines will.have been impl~mented. At present two t,pes of inactivated vaccine a~e 1n use, one der1ved from mouse brain and the other from primary hamster k1dney cells. The mouae brain vaccine is highly purified well-defined and has been adopted for production in leveral countries.' Japanese . , encephalitis virus strains appear to be clustered into two antigenic ~roups. The inclusion of one strain of virus from each of these ~roups in the vaccine has improved its potency in the leboratory. From the inforaation presented at this meeting, it .eeas that high priority will have to be ~iven to standardization of potency tests so that vaccines can be compared directly. The rapid and early detection of infection witb Japanese encephalitis viruses has baen successfully accomplisbed uain~ an ELISA test based on IgM antibody capture techniques. The work on development of second generation vaccines is in pro~ress. Data on the analysis of the topography of the envelope protein V3 su~.ested thst there were eight domains of antigenic determinants on the protein. Preliminary data was also presented on cloning the genome of JE virus. 4.
DENGUE HAEMORRHAGIC FEVER
Den RUe fever and dengue haemorrhagic fever (DHF)/ahoek syndrome are serious problems in south-east Asis and the Pacific re~io.s. In receDt yeara, the viruses have spread and the epidemic of DHF in Cuba in 1981 illustrated the potential for serious outbreaks in areas previously fr.e of DHF. The developeent of live attenuated vaccine. has been pursued by tvo different research or~anizations. Followin~ different basic strategies in the selection of vaccine strains and attenuation of them, each appears to have a candidate vaccine for dengue 2. In a recent trial study of one vaccine, the preliminary analysis of the results suggests that the vaccine will be immunogenic. The immunity generated in volunteers needs to be followed for longer periods. Another dengue 2 vaccine produced only transient, low level antibody in volunteers not previous 1, immune to ,ellow fever virus. No attempts have been made to boost w .... in~ ilBllUnity. The problems presented by the development of polyvalent vaccines remain. Candidate vaccines are currently beiq~ developed for tbe other serotypes and the formulation and administration of a vaccine containin~ four live sttenuated viruses may present future problems. Rapid and early diagnosis of denRue appears to be mo~e pr08isin~, techniques based on detection of virus. The IgM capture ELISA technique used so successfully for JE v4rus requires furtker testing for denRue virus and other methods need to be explored. The availability of cDNA probes for use in ~ ~ hybridization tests will iDCrease as work on the molecular analysis of dengue virus genome. proceeds. As with Japanese encephalitis virus, there is work in progress on recombinant DNA vaccines but lack of fundamental knowled~e of the structure of the virus ~enomes hinders prORress in this area of research. usin~
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5. 5.1. Vaccines
RECOMMENDATIONS
5.1.1 At present no vaccine exists for haemorrha~ic fever with renal syndrome (HFRS). WHO should encoura~e research in vaccine development. Research on inactivated Hantaan virus anti~en is su~~ested as a first step in the development of an HFRS vaccine. 5.1.2 Since no ~ood animal model reflectin, HFRS exists, WHO should encouraRe research in model development. 5.1.3 Inactivated Japanese encephalitis (JE) vaccines are prepsred in China, Japan, and the Republic of Korea. WHO has recommended standardization of inactivated vaccines. The Group stressed the need for WHO to encourage the transfer of techniques for conducting potency and safety tests. 5.2. Vaccine development by modern biotechnology 5.2.1 WHO should support and encouraRe research on the development of second Reneration Japanese encephalitis (JE) vaccines utilizing modern techniques of molecular biotechnology. 5.2.2 WHO should support the application of recombinant DNA techno lORY to identify Renetic markers of denRue virus attenuation and alternative methods for vaccine production. 5.3. Laboratory diaRnosis 5.3.1 WHO should encourage development of standard dia~ostic techniques for HFRS in order to compare results obtained in the different laboratories. 5.3.2 WHO should coordinate the preparation and distribution of panels of reference sers for comparative IgM antibody capture-ELISA testing for Japanese encephalitis. These reference sera should be evaluated by three laboratories prior to final circulation. 5.3.3 WHO should continue to support the development of rapid diagnostic methods that can be applied in the earliest phase of dengue infections. Ideally, these should identify the dengue serotype and be applicable in areas where other flaviviruses co-exist. Assays for viral antigen specific antibodies, or virus-antibody complexes should be considered. Research is needed on the development of cDNA probes for the detection of denRue nucleic acid in infected cells and viraemic blood. 5.3.4 Because of differin~ success between laboratoriea in the use of the I~M-ELISA for den~ue, WHO should coordinate collaborative studies usin~ this technique by exchan~in~ coded sera, anti~ens and test procedures for comparative evaluation.
5/6
5.3.5 Laboratories currently employing intrathoracic inoculation of adult Toxorhynchites splendens for dengue virus isolation should attempt the intracerebral inoculation of mosquito larvae, to compare the sensitivity and ease of performance. WHO should coordinate the training of laboratory personnel in larval mosquito inoculation. 5.4. Epidemiolosy and surveillance 5.4.1 Japanese encephalitis virus isolates from different countries together with their immune sera should be sent to the WHO Collaboratina Centre for Reference and Research on Arboviruses in the National Institute of Health, Japan. These should be accompanied with the history of isolation to be recorded on a standard form. The different virus strains should be made available to other interested and competent laboratories. 5.4.2 Certain surveillance pro~rammes are encouraged. Epidemiolo~ical studies should evaluate risk factors for dengue haemorrhagic fever in order to assess the applicability of monovalent vaccines in some areas. Locations in which Phase I and II testinl of candidate vaccines is done must be regularly monitored for natural transmission of denlue and other flaviviruses. 5.4.3 WHO should provide support to countries wishinl to test candidate dengue vaccines. 5.S Research on pathophysiology
5.5.1 Basic research on the pathophysiology of dengue haemorrhagic fever is encouraled. Specific aims should include the identification of biochemical factors which are clinically useful for predicting the onset of shock; 5.6. Newsletter 5.6.1 The ~reatly increased number of recent developments in HFRS research makes the rapid and efficient exchange of new data between specialists absolutely essential. It is important that a newsletter be inaugurated, to be distributed at intervals to interested HFRS investigators. It is further suggested that the WHO Regional Office for the Western Pacific should cooperate with collaborating centres and interested scientists in the preparation and dissemination of such a Newsletter. 5.6.2 The WHO Regional Office for the Western Pacific should cooperate with collaboratinl centres and interested scientists in the preparation and dissemination of a newsletter on Japanese encephalitis virus research.
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ANNEX 1
AGENDA
Tuesday. 28 AUIust 0800 0830 0900 Registration Opening Ceremony Epidemiolo~y.
and
rea~ents
diagnostic methods of HFRS Dr H.W. Lee Dr T. Kitamura Dr N.Hashimoto
Republic of Korea Japan USA
1015 1030
Coffee break Characterization of Hantaan virus and development of vaccine Republic of Korea Japan USA Dr H.W. Lee Dr T. Kitamura Dr M. Takahashi Dr J. Dalrymple
1200 1330
Lunch break Rapid diagnostic methods for JE (12M capture method) Coffee break JE vaccine Quality control of vaccine China Thailand Bivalent vaccine
Dr A. Igarashi Dr C. Hoke
1415 1430
Dr Li Ho-min Dr S. Nhadirat Dr K. Fukai Dr A. Ova
Quality of JE vaccine
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Annex 2
Wednesday, 29 0900
Au~ust
Rapid
dia~nosis
of
den~ue
Dr T. Pan£ nr C. Hoke Dr A. IS!,arashi
1000 1015 1045
Coffee break Human trial with attenuated vaccine Anti~enic
Dr Natth Dr Bancroft
analysis of den~ue virus and development of component Dr J. Blok
vaccine
1200 1400
Lunch break Standardization of dia~nostic methods and rea£ents HFRS Den~ue
JE
Thursday, 30 0900
Au~ust
Group discussions
HFRS Den~ue
JE 1000
Coffee break Group discussions (cont'd.) Lunch break Drafting of recommendations
1015 1200 1400
Friday, 31 August 0900
Finalization of recommendations
1200
Closing ceremony
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ANNEX 2
LIST OF MEMBERS, CONSULTANTS, OBSERVERS AND SECRETARIAT 1. MEMBERS
Dr Janet B10k Virologist Queensland Institute of Medical Research Herston, Brisbane Queensland 4006 Australia Dr Barry Gorman Senior Virologist/ Principal Research Fellow Queensland Institute of Medical Research Herston, Brisbane Queensland 4006 Australia Dr Guido van der Groen Head of Virology Department Institute of Tropical Medicine Nationa1estraat 155 B-2000 Antwerpen Belgium Dr Nobuo Hashimoto Department of Veterinary Public Health Facultv of Veterinary Science Hokkaido University Nishi 9-chome, Kita 18-Jo Kita-ku, Sapporo Japan Dr Akira Igaraehi Professor Department of Virology Institute for Tropical Medicine Nagasaki University 12-4 Sakamoto-. .chi Nagasaki Japan
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Annex 2
Dr Kyong Ho Kim Diretor Korea Green Cross Research Institute
729, Seocho-Don2, KangNam-Ku Seoul T-135 Republic of Korea Dr Takashi Kitamura Chief, Division of Poxviruses and Special Pathogens National Institute of Health Muravama Annex Nakata, Musashimurayama Tokyo 190-12 Japan Professor Ho Wang Lee Dean
Institute for Viral Diseases Korea University Medical Col1e~e 1 Anamdon~ Seonghuk-ku Seoul Republic of Korea Or Li He-min National Institute for the Control of Pharmaceutical and Biological Products Beijinl!, People's Republic of China Or Akira Ova Director Department of Virology and Rickettsiology National Institute of Health 2-10-35, Kamiosaki, Shinal!awa Tokyo 141 Japan
I, Dr Sung Bok Paik Director Department of Virology National Institute of Health Nokbun-dong, Eunpyung-~u Seoul 122 Republic of Korea II
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Annex 2
Dr Tikki PanR Associate Professor Department of Medical Microbiolo.V Faculty of Medicine University of Mslaya Kuala LU1IIpur Malaysia Dr Michiaki Takahashi Director and Professor Research Institute for Microbial Diseases Osaka University 3-1. Yamadaoka. Suita Osaka Japan Dr U.T. Vitarana Medical Research Institute Baseline Road Colombo 08 sri Lanka Dr Kotaro Yasui Department of MicrobioloRy Tokyo Metropolitan Institute of Neurosciences 2-6 Musaahidai. Fuchu Tokyo Japan
2.
CONSULTANTS
Professor Natth Bhamaraprayati Rector Mahidol University Bangkok Thailand Dr Konosuke Fukai Chairman. Board of Directors Research Foundation for Microbial Diseases Osaka University 3-1 Yamada-Oka Suita. Osaka Japan
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Dr Nakao Ishida Professor and Chairman Department of BacteriolollY Tohoku University School of Medicine 2-1 Seiryomachi Sendai 980 Japan Dr K. Pavri Director National Institute of Virology Pune India
3.
OBSERVERS
Dr William Bancroft Chief, Department of Virus Diseases Walter Reed Army Research Institute Washinaton, D.C. 20307 United States of America Dr Joel Dalrymple United States Army Research Institute of Infectious Diaeases (VirolOI\Y) Fort Dedrick Fredrick, Msryland United States of America Dr Charles H. Hoke Chief, Department of Virology Armed Forces Research Institute of Medical Sciences Ban!\kok Thailand Dr Nobuo Koinuma
Deputy Director International Affairs Division Ministry of Health and Welfare 1-2-2, Kasumi!\aseki Chivoda-ku Tokyo Japan
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Annel< 2
Dr Nadhirat Sangkawibha Director-General Department of Medical Sciences Yod-se, Bangkok 10100 Thailand
4.
SECRETARIAT
Dr Taguir Bektimirov Chief Medical Officer Virus Diseases Unit WHO Headquarters Geneva Dr Lee S. Self Scientist Vector Biology and Control WHO Regional Office for the Western Pacific Manila Dr Akira Shirai Medical Of Hcer Tropical Diseases Research WHO Regional Office for the Western Pacific Manila Dr Taku8ei Umenai Re~ional Adviser in Communicable Diseases WHO Regional Office for the Western Pacific Manila
15 ANNEX 3
TECHNICAL BASIS FOR RECOMMENDATION ON HAEMORRHAGIC FEVER WITH RENAL SYNDROME Introduction Haemorrha~ic fever with renal syndrome (BFRS) is a public health problem throughout most of the European and Aaian land mass. Althou~h predominant Iv associated with rural areas, it is now bein~ recognized as an urban problem in some countries, and also preaents a particular hazard to laboratory staff who use rodents for biomedical research. In wild rodents (rats, mice and voles) the infection is asymptomatic. Human infection with the HFRS agent(s) is sporadic, but under special circumstances epidemics occur; the infection may be completely silent, or associated with mild or severe disease. Severe cases are usually seen in east Asia. The epidemiological features of the disease vary from country to country and depend upon a variety of factors, the elucidation of which requires a multidisciplinary approach. Recently, not only Hantaan virus, the etiological a~ent of HFRS in Asia, but also Puumala virus, the possible etiological agent of Nephropathia epidemica in Finland, were isolated in Vero-E6 cell culture. In addition, more than 30 other Hantaan-related viruses have been isolated in Vero-E6 cell culture. Eight viruses studied by RNA sequencing have been shown to have a common 3' nucleotide sequence different from those of existing genera in the Bunyaviridae family. This su~~ests the existence of a possible new genus in the Bunyaviridae family which includes this new group of Hantaan-related viruses.
With the continuing isolation of new vir~ strains, surveillance becomes very important and further research on these viruses is needed to identify the common antigenic determinants and to determine strain differencps. This will be most helpful in determining the best strstegy for future vaccine development. Recommendations 1. EpidemiOlogy and surveillance
It is necessary to establish global surveillance for this disease, provide seroloRical confirmation of suspected cases, define geographical areas of endemic Rantasn virus infection as well a. provide the relevant data necessary to define apparent to inapparent infection ratios and incidence-prevalence data for areas with known disease. 2. Risk ~roup9 ~roups
Existin~ epidemiological information su~gests the followin~ individuals will be at risk and will be the first candidates for vaccination:
or
2.1
people living or travellin~ in endemic areas where the severe form of the disease is known to occur;
16 Annex 3
2.2 2.3 2.4
laboratory workers exposed to experimental and natural HFRS infection; personnel of animal breeding rooms; farmers, foresters, soldiers, soil testers, and construction workers in endemic areas and veterinarians in contact
with infected animals. 3. Research priorities for vaccine development 3.1 Amon~
the many problems facin~ the development of any vaccine for HFRS is the absence of a ~ood animal model for virus vaccine testin~. The best animal model for this disease would be an animal infection that closely mimics human HFRS disease. It is proposed that the team of experts attempt to standardize the criteria for a classification of disease severity which can be uniformly interpreted and applied throughout the world. Such a classification would require a first-hand comparison of the clinical forms of the disease in the different ~eographical areas reportin~ varying signs and symptoms. This standardized disease classification would provide a basis for animal patho~enesis studies attemptin~ to provide a suitable animal model of human disease and provide insi~ht into pathogenic mechanisms. As a part of the classification of the disease, every effort should be made to isolate viruses from clinical cases of varying severity. Such isolates would allow the detailed comparison of antigenic differences and virulence markers if they exist and can be correlated with disease severity. Pendina the development of a suitable animal model, virus isolates obtained could be examined for variation in patho~enicity, antigenicity and immuuogenicity in mice. The potential virus vaccine candidate must contain the important immunogenic determinants necessary to immunize and protect against all strains currently capable of causing severe disease. It is evident that the worldwide surveillance of the disease as well as the evaluation of future candidate vaccines rely entirely on reliable methods for the detection of anti~en, viral infectivity and specific antibodies. Priority should therefore be given to developing techniques which Rive reproducible results and can be standardized in order to compare the results obtained in different collaborating laboratories (See Appendix).
3.2
3.3
4.
Approaches to HFRS vaccine development 1••
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Development of live attenuated virus vaccines Based on the limited information currently available, attenuated virus vaccines for HFRS cannot be recommended as a major objective of the HFRS vaccine research plan. At present, there is no means of assessing attenuation of any candidate virus
preparation.
Similarlv, the propensity of these viruses to infect
17/IR Annex 1
persistently both animals and cell cultures raises serious concern about the possibility of virus persistence in vaccine recipients. The demonstration of a se~mented genome ~reatly increases the probability of virus vaccine reassortment with other wild type viruse. circulating in nature. For the reasons stated, the development of a live attenuated vaccine cannot be recommended at the present time. 4.2 Development of inactivated or sub-unit vaccines Research on inactivated Rantaan virus antigens is su~~ested as a first step in the development of a RFRS vaccine. Althou~h immunogenicity and the induction of hi~h titred humoral antibody are important for the future success of any HFRS vaccine, protection against aerosol challen~e as well as parenteral inoculation must be considered because of currently established mechanisms of transmission. 4.2.1 The possibility that virus anti~en can persist in animal tissue in the presence of hi~h titred circulating antibody must also be considered in evaluatin~ vaccine efficacy. Antigen elimination should be a criterion for successful animal immunization. Sub-unit vaccines composed of major virus immuno~enic proteins should be considered in vaccine development reaearch 'oth aa a potential means for increased immunogenicity and as an aid in the identification of important Rantaan-related virus antigens.
4.2.2
5.
Vaccination of laborptory animals Rantaan related virus infection of laboratory rodents remains a potential source of human disease and needs investi~ation. The worldwide prevalence of laboratory infection is not yet known but should be determined by coordinatin~ the diagnostic capabilities of the WHO Collaborating Centres for Reference and Research on Raemorrhagic Fever with Renal Syndrome (RFRS) to increase the animal serological surveillance capabilities of laboratory animal breeders. In the event that control measures such as caesarian section and foster nursing do not turn out to be efficient means of virus eradication, the use of an animal vaccine must be considered.
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APPENDIX
1.
Antibody
testin~
For lI;eneral human or animal serosurveys, the method in Ileneral use is an indirect immunofluorescent test with E6 clone of Vero cells infected with the strain 76-118 of Rantaan virus. This test has been shown to detect antibody which mayor may not react with other Hantaan strains. The antigen slides are availa~le from several WHO collaboratin~ laboratories. There i. a need to evaluate carefully the sensitivity and specificity of the several available serolOllical tests for HFRS virus antibody, includinll; IFA, ELISA an4 lARA. Theae must all be eompared with neutralization tests. The evaluation of these testa viII be carried out in laboratories in Beoul, Tokyo, Osaka, Antwerp, Moscow, Bethesda and Atlanta. A standard plaque reduetion neutralisation test protocol will be distributed to all of tbe laboratories with a penel of coded antisera from various lleOllraphic areas in the world. Protocols for the IFA, ELISA, CF and lARA will also be distributed. The 8ame three strains of virus will be used in each laboratory (76-118; Puumula; and urban rat strain). Data will be evaluated from all of the laboratories by Dr Ho Wang Lee, who is the coordinator of this study. 2. Rapid diagnosis of HFRS
Based on data from China and the Republic of Korea, it appears that IIl;M antibody sccurs very early in RFRS infection such that almost 100% of patients are positive at day 3 of the illness. The use of IFA test with specifie I~M antibody is therefore recommended for current use as a rapid diallnostic technique. More thorough evaluation of this technique is needed, and its reliability in various laboratories should be evaluated. Thus, a panel of coded acute sera from HFRS patiants will be prepared and sent to collaborating laboratories for ~omparative evaluation of the IFA and~ chain capture ELISA techniques usin~ strain 76-118 as standard anti~en. Dr He Wang Lee, Direetor of the WHO Collaboratin~ Laboratory in Seoul, will prepare this panel of antisera for evaluation by the participatinll; laboratories. Protocols for carryin~ out the two tests will be distributed with the sera. The data .ill be collated and evaluated by Dr Lee and re-distributed to WHO and the p.rticipatin~ laboratories for comment. 3. It i. recommended that the monoclonal antibodies developed a~ainst HFRS should be used as sosn as posBible to help to differentiate smon~ the incressin~ n~ber of HFRS virus strains actually known. Therefore, monoclonal antibodies supplied by Dr J. McCormick and Dr Yamanishi will be distributed a.on~ CDC, Fort Detrick (USA), Korea, Tokyo, Osaka, USSR, Scandinavia aud Bel_ian laboratories in order to test the specific reaction patterns on all HFRS strains so far available in the E-6 cell line. The data will be collated and evaluated at the Institute of Tropical Medicine in Antwerp. A standard protocol for testin~ will be sent to all contributors of the pro~ral1lllle.
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Appendix
4.
"FRS - aDtigen detection in the field studies
Information of the prevalence of HFRS related virus anti~en amon~ rodents csptured in a desillnated locus may be vital in assessinl! the epidemiolo,!ical situation of that locus. There are three methods used successfully in participatiril! laboratories, i.e. IF of tissue sections, ELISA and IAHA. These methods should be compared by the use of standardized antisera a~ainst representative strains of HFRS virus supplied by the Tokyo coordinatin,! laboratory, with special reference to efficacy of antillen detection, simplicity of the facilities required, rapidity and ease of performance and reproducibility of the results under various field conditions. A set of monoclonal antibodies, includinll both cross-reactive and type-specific react9rs, could be most useful. Standsrd methods and procedures mu.t be .uitable for application under field conditions.
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