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Competing Interests: There are no conflicts of interest.

Competing Interests: There are no conflicts of interest.

Substantial progress has been made towards achieving global eradication of poliomyelitis by the end of the year 2000; the goal set by the World Health Assembly in May 1988. The basic strategies to eradicate polio are: attaining high routine coverage with at least three doses of OPV; conducting national immunization days (NIDs) in polio endemic countries; establishing a sensitive system of acute flaccid paralysis (AFP) surveillance to track wild poliovirus circulation; and conducting "mopping-up" immunization when polio is reduced to focal transmission. By the end of 2000. India was in the midst of the sixth National Immunization Days (NIDs). Surveillance system for Acute Flaccid Paralysis (AFP) continued to achieve the recommended non-polio AFP rate of at least 1 per 100,000 population < 15 years per year (1.88 for week 51 ending 23rd December 2000), the adequate stool specimen collection rate was 83% that met the target of > 80%. Some States in the south and west have started to implement mopping-up immunization campaigns as the end-stage strategy to eliminate the last remaining foci of transmission. While most of India appears to be well placed to eradicate polio by the end of 2000 or shortly thereafter, concerns remain about low coverage in parts of the densely populated northern States of Uttar Pradesh and Bihar where high intensity transmission persists. The Government of India has embarked upon an intensified strategy that relies on extra rounds of NIDs; house-to-house immunization to reach previously missed children; and aggressive mopping-up campaigns including pre-emptive mopping-up in the known reservoirs in Uttar Pradesh and Bihar. Extensive microplanning and supervision of the supplementary immunization activities is critical to achieve the target of polio eradication.

A Comparative study of three types of measles vaccines was undertaken among 1005 children. Of these 527 were vaccinated with the Serum Institute of India (SII) vaccine, 230 with Schwarz (SC) and 248 were vaccinated with Edmonston-Zegreb (EZ) vaccine (imported from Zegreb). Though the majority of children reacted favourably with all the three vaccines (SII: 98.43%; SC: 93.40%; EZ: 93.0%) with a rise in titre, but the percentage of seroconversion was significantly higher with the SII vaccine (p < 0.01). The Schwarz and Edmonston Zagreb vaccines showed significantly less GM titre as compared with the other age group i.e. 9-12 months (p < 0.05). With Serum Institute of India (SII) vaccine the GM titres were almost similar in the different age groups. The overall GM titre obtained with the SII vaccine was significantly higher than the SC vaccine (p > 0.001) as well as the EZ vaccine (p > 0.001). It is of interest to note that among the infants, 22.5% children had measles antibody in them before vaccination.

Two proteins (putative receptors) of 60 and 38 kDa, for chikungunya (CHIK) virus were detected in the brush border membrane fraction (BBMF) of the normal population of Aedes aegypti mosquitoes. Mosquitoes were infected orally with CHIK virus and infectivity checked by testing the head squashes. BBMF was prepared from proved positive and negative mosquitoes. The receptor proteins were found to be present in both the proved genotypes. However, dot-b'ot assays showed that the CHIK virus binding activity of BBMF/mg protein was noticeably low in the proved negative mosquitoes as compared to the positives. BBMF from the larvae of the normal populations also showed the presence of the receptor proteins, binding to CHIK virus. Receptor proteins from larvae as well as the adults were found glycosylated. CHIK virus receptor proteins of 24, 45, 58, 60 and 62 kDa were also seen in the membrane fraction of the C6/36 cells.

There has been a growing interest during recent years in the role of free radicals and lipid-peroxidation at tissue-level for the causation of cancer and other age-related diseases like atherosclerosis, rheumatoid arthritis, cataract etc. Free radicals and increased lipid peroxidation play a significant role for causation of human diseases by oxidative damage and functional degeneration of the tissues. Vitamin C, a well-known dietary antioxidant, and other enzymatic antioxidants like glutathione can protect the lipids of lipoproteins and other biomembranes against peroxidative damage by intercepting oxidants before they can attack the tissues. But cigarette smoking was found to affect the antioxidant protective action of Vitamin C, glutathione etc. A group of adult male smokers in this study were found to have lowered Vitamin 'C' & glutathione levels, but increased lipid-peroxide levels in their blood. Thus the increased pathogenicity of the smoking may also be due to indirect biochemical effect of enhanced oxidative stress by increased lipid-peroxidation and lowered Vitamin C & other antioxidants at tissue-level.

During 1992-96, outbreaks of buffalopox zoonosis were reported from different villages in Jalgaon, Dhule and Beed districts of Maharashtra State. In humans, pox lesions were observed on the hands whereas in affected buffaloes and cows the lesions were noticed mainly on the teats and udder. Twenty two virus strains were isolated from the skin scabs collected from infected humans and milch animals. Neutralizing antibodies were detected not only in the sera of affected humans but also in their contacts. Detection of antibodies in young individuals from endemic area, who were neither vaccinated for smallpox nor had any contact with buffaloes or history of any poxvirus disease, is suggestive of occurrence of subclinical infection. A few children who had no contact with infected animals also showed clinical manifestations with disseminated lesions on the face, arm and buttocks, and thus suspected to have acquired infection through their infected parents or other family members indicating a possible man to man transmission. Therefore, in the light of discontinuation of smallpox vaccination, buffalopox outbreaks need to be monitored carefully as this may emerge as a serious zoonotic disease in India.

Experimental studies were carried out to determine the vector potential of four species of mosquitoes to West Nile (WN) virus, viz. Culex tritaeniorhynchus, Cx. vishnui, Cx. bitaeniorhynchus and Cx. univittatus. All the four species of mosquitoes successfully transmitted and supported the growth of WN virus. The study indicated that the four species of mosquitoes could act as potential vectors of WN virus in nature.

An emerging viral infection may be a totally new disease with undescribed symptomatology as it was in the case of Kyasanur forest disease in Karnataka, but more often it is an introduction of a known or little known disease in an area where the disease did not occur earlier e.g. yellow fever in Kenya or Rift valley fever in Egypt. The virus may show altered degree of virulence due to many changing factors as in the case of the different haemorrhagic fevers. Many factors may contribute to the emergence of viral infections which may be genetic exchanges or mutations; adaptation to new hosts or vectors; and changed social patterns of humans like urbanization, rapid transport, trade, migration of people or of vectors, strain on civic facilities or changing moral values and life-styles. Large scale changes in ecology due to global warming, deforestation or afforestation, building of dams or canals, changed agricultural practices, rearing of livestock or birds may also contribute to emergence of viral diseases. A number of emergent virus infections relatively important to India have been discussed. To combat emergent virus infections, a comprehensive strategy needs to be evolved. A national viral surveillance system needs to be established. Epidemiology of virus diseases needs to be studied in depth. Development of diagnostic reagents and their supply to investigating centres, a Central serum bank, and a virus respository are important factors. Research and development on viruses, as regards the epidemiology, diagnosis, pathogenesis and vaccinology of virus infections need to be strengthened. An international network of databases of virus infections needs to be instituted. A global network for the diagnosis and containment of emerging viral diseases is advocated.

AN overview of the general principles of counselling for HIV/AIDS are overviewed. Relevance of 'shared confidentiality' in Indian context is recommended.

Nearly 2000 serum samples collected from different risk groups from Pune and Bombay metropolitan areas were tested for antibodies to hepatitis C virus (anti-HCV) by Recombinant Immunoblot Assay-3 (RIBA-3). Patients undergoing haemodialysis showed 24.5 per cent seropositivity whereas 5.7 and 5.3 per cent of multiply transfused patients (>2 units) and chronic liver disease patients respectively were anti-HCV positive. Leprosy patients had almost 0.7 per cent seropositivity. In other risk groups the positivity rate was nil. In normal population only one out of 830 persons had anti-HCV antibodies. It is therefore apparent that the prevalence of hepatitis C virus (HCV) in western India is not high. However, special care needs to be taken for dialysis patients. As none of the 430 pregnant women and 86 children below the age of 5 yr were anti-HCV positive, vertical mode of HCV transmission seems to be negligible.

IgG and IgM class of antibodies to measles virus were estimated in the plasma samples of 211 unvaccinated infants living under overcrowded conditions in Pune, Maharashtra. IgG antibodies to measles virus, in high titres, were detected in 52.1 per cent and IgM antibodies in 31.3 per cent of the infants studied. Most of the infants with IgM antibodies to measles virus had not suffered from an attack of measles. A history of contact with an older child with overt measles was available for 13.3 per cent of the 211 infants studied. Among these infants with such a contact history, 82.1 per cent had IgM antibodies to measles virus. These observations suggest that many infants studied by us, had experienced subclinical measles infections and many times older children with measles at home or in the neighbourhood might have transmitted such infections.

Anti-hepatitis A virus IgM capture ELISA was developed by using the reagents produced in the NIV laboratory. The major reagents of the assay were anti-human IgM antibody, hepatitis A virus (HAV) and anti-HAV IgG-horse radish peroxidase (HRP) conjugate. Of these, anti-human IgM antibodies were generated in rabbit against IgM secreted by human hybridoma clone(G3). HAV was derived from buffalo green money kidney cell line infected with HM-175 strain. Virus purified from the cell lysates was used for immunization of rabbits and guinea-pigs. There was very low anti-HAV response. A seropositive rhesus monkey was inoculated with monkey adapted strain of HAV to boost the anti-HAV antibody titre. Anti-HAV IgGs derived from hyperimmune sera of monkey and hepatitis A patient were conjugated with HRP. The preparations of conjugate--particularly human antibody--HRP conjugate yielded highly satisfactory results in anti-HAV capture ELISA. The assay appears to be specific, sensitive and quick and is useful in differentiating acute HAV infection from other acute infections caused by B, E and non-A non-B hepatitis viruses.

A formalin inactivated Kyasanur forest disease (KFD) virus tissue culture vaccine produced by the health department of the State Government of Karnataka at Shimoga was administered in Shimoga, Uttar Kannada and Chikmangalur districts during 1990-92 KFD epidemic seasons. The selection of places for vaccination was based on the prevalence of KFD activity in previous years; villages adjacent to KFD affected areas and the villages from which mortality in monkeys was reported. A total of 284 villages was covered under vaccination; 26850 individuals received one dose whereas, 61302 received two doses of vaccine. No untoward reaction was observed in any of the vaccinees. In the 72 KFD affected villages there were 14 patients among 9072 and 10 among 21083 vaccinees receiving one and two doses respectively, whereas 325 patients were reported among 37373 unvaccinated individuals during the same period. In 1990-91 the number of males patients was more than females whereas, in 1991-92 the ratio was reserved. On analysis indicated that the vaccine has a highly significant protective effect.

Studies were carried out to determine the effect of prolongation of incubation periods, cocultivation with normal buffalo green monkey kidney (BGMK) cells and different concentrations of foetal calf serum (FCS) on the production of hepatitis A virus (HAV) by BGMK cell line persistently infected with HAV strain HM175. HAV could be detected from week 1 onwards. However, maintenance of cultures beyond this period was found to yield substantially higher quantities of virus. Cocultivation of persistently infected cells with normal BGMK cells also improved the antigen yields. Different concentrations of FCS did not show any effect on the amount of virus produced. The cell line was maintained up to 46 passages during which there was continuous production of HAV in the cells and release of small amounts of virus in the culture supernatants. Cell associated and cell free viral particles were found to be infectious. Supernatant derived virus was a highly suitable inoculum for infecting other susceptible cell lines. Persistently infected BGMK cell line appears to be a reliable and economical source to derive HAV in adequate amounts for diagnostic and research purposes.

Serum thyroxine (T4), triiodothyronine (T3), thyroid stimulating hormone (TSH) and thiocyanate (SCN) levels in 40 smokers smoking filter or non-filter cigarettes were compared to those in 20 control subjects. Both types of cigarette smoking resulted in a decrease in T4 (P < 0.01) and increase in TSH (P < 0.05) to the same degree as compared to control. T3 was found to be higher in the smokers but the rise was not significant. SCN was elevated significantly in the smokers. The filter attached to the cigarette was found to have little or no effect on the alteration of the levels of the biochemical indices studied.

Following the reports of epidemics of febrile illness from several rural and urban areas of Gujarat state (India) in 1988, epidemiological investigations were carried out and dengue (DEN) virus activity was demonstrated in large cities such as Surat and Rajkot as well as several villages in Sabarkantha district. Two strains of dengue type-2 each were isolated from human sera from Surat city and a village in Sabarkantha district. Six strains of dengue virus were isolated from Aedes aegypti mosquitoes collected at Chotasan village, two of which were confirmed as DEN type-2. Of the 560 patients' sera tested from different areas (including villages and townships), 122 showed evidence of dengue infection and another 236 showed a broader reaction with flaviviruses. Entomological investigations showed a widespread distribution of Ae. aegypti both in urban and rural areas. In the household conditions this mosquito was found to breed predominantly in containers with non-potable water. Amongst these, cement containers manufactured in towns and distributed to the villages seem to play an important role in the spread of this species. In non-residential areas prolific breeding of Ae. aegypti was observed in automobile tyre dumps, and varied types of scrap, in towns and villages. Distribution and relative prevalence of the species were studied in 46 towns and villages, covering the spectrum of rural-urban-continuum. These studies provide an indication regarding the mechanism of the spread of DEN virus through peoples' movement, transport, the process of urbanisation etc.

Ae. aegypti mosquitoes from rural areas of Maharashtra state were subjected to insecticide bio-assays. All the populations showed resistance to DDT at larval and adult stages. No resistance to deltamithrin and malathion was detected at any stage. Biochemical analysis of these mosquito populations showed that resistance to DDT was probably due to increase in the kinetics of glutathione S-transferase.

Ten non-pregnant female monkeys and four pregnant monkeys (all Macaca mulatta) in the last third of their gestation period were infected intravenously with the stool sample of a patient with hepatitis E virus infection (immuno-electronmicroscopy positive for hepatitis E virus). Four more non-pregnant monkeys were inoculated with a lower dose (less number of virus particles by IEM) of a stool sample collected on a different day from the same patient. The average incubation period as evidenced by the rise of serum alanine transferase in the non-pregnant monkeys, was 36.4 +/- 4.9 days. The dose of the virus did not affect the incubation period. Two of the pregnant monkeys had incubation periods of 9 and 13 days respectively. They delivered healthy babies on 40th and 53rd day respectively after inoculation. At the age of 11 months, both babies were negative for anti-HEV antibodies. One monkey which delivered a healthy baby on the 2nd day after inoculation had incubation period of 36 days. The baby of this monkey was anti-HEV positive at the age of 11 months. The incubation period was 41 days in the fourth monkey which delivered a macerated foetus on the 36th day after infection. No fatality was recorded in the infected monkeys. Bile samples collected from all monkeys showed strong signals in nested polymerase chain reaction (PCR). It seems that the incubation period in pregnant monkeys was determined by the state of pregnancy.

Susceptibility status of five populations of Ae. albopictus mosquitoes from Maharashtra state, to DDT, malathion, fenitrothion, bromophos, propoxur and deltamethrin was studied and compared with the laboratory population. Four populations survived when adults were exposed to 4 per cent DDT impregnated paper for 2 h; though three of these populations had lower DDT-LC50 values at larval stages in comparison with the laboratory population. Results of topical application of DDT on these four populations supported these findings by showing comparatively higher LD50 values at adult stages in comparison with the laboratory population. All the populations were highly susceptible to other pesticides tested i.e., malathion, fenitrothion, bromophos, propoxur and deltamethrin. These populations were distinguished from each other by esterase isoenzyme patterns.

Two hundred serum samples were tested to detect the presence of HIV-2 infection in Maharashtra state. The serum samples were derived from various groups including those with high risk behaviour for HIV infection. All samples were tested by three combined HIV-1 and HIV-2 ELISA kits. The reactivity was confirmed by LiaTek HIV 1+2 immunoblot. The study confirmed that HIV-2 infection exists in Maharashtra, as 14 samples showed antibodies to HIV-2 and 14 showed antibodies to both HIV-1 and HIV-2. Antibodies to HIV-2 or HIV-1 and HIV-2 were detected mainly in persons with high risk behaviour.

A highly sensitive polymerase chain reaction (PCR)-based diagnostic assay was developed to detect the presence of hepatitis B virus (HBV) in human serum. The assay involved amplification of HBV DNA sequences using primers specific to HBV. This in vitro enzymatic amplification technique, when used in combination with molecular hybridization assay can detect HBV genome in the patient's serum, with a higher degree of sensitivity than achieved with dot blot assay. The assay can identify samples containing 3-10 virus particles.

As beta 2 microglobulin (B2M) has been found to be elevated in immunological disorders and also in HIV infection, its levels were studied in 475 HIV seropositive, asymptomatic persons; 101 HIV seronegative persons from high risk groups for HIV and 99 healthy controls. The B2M levels in asymptomatic HIV seropositives are found to be significantly higher than healthy controls (1.0 mg/1 to 2.7 mg/1, P less than 0.001) and HIV seronegatives from high risk groups (1.1 mg/1 to 2.7 mg/1, P less than 0.001). Two hundred and thirty four (49.26%) seropositives showed increased levels of serum B2M. Thus, quantitative analysis of B2M may be useful as an early nonspecific marker of HIV infection and immune dysfunction. The prognostic value of B2M was assessed in a follow up study of 54 HIV seropositives in a 2 yr period. Within this period, B2M levels were found to be significantly increased in these subjects (1.2 mg/1 to 4.6 mg/1, P less than 0.001). Three of the subjects who showed high increase in the B2M levels, progressed to AIDS-related condition, whereas one progressed to persistent generalised lymphadenopathy. Thus, the rising levels of B2M appears to correlate well with disease progression.

During the Japanese encephalitis (JE) epidemic in 1988 at Gorakhpur, Uttar Pradesh, 34 cerebrospinal fluid (CSF) samples with 16 matching sera from 34 anti JEV IgM positive (confirmed JE) and 24 CSF samples with 4 matching sera from 24 anti JEV IgM negative (clinical encephalitis) patients were collected and tested for presence of JEV specific IgG by ELISA. Eighteen CSF samples and 8 matching sera from confirmed JE and 5 CSF samples and one matching serum from clinical encephalitis patients positive for JEV specific IgG were further assayed for subclass specificity using specific murine monoclonal antibodies. Almost all the samples exhibited IgG1 as the virus specific subclass. In addition to IgG1, one serum and one CSF sample each from two different confirmed JE patients showed the presence of virus specific IgG4 and IgG3 respectively. Half of the confirmed JE and clinical encephalitis patients exhibited intrathecal synthesis as evident from either elevated IgG index or CSF IgG/CSF albumin ratio. Most of the patients who recovered had predominantly virus specific IgG1 in CSF. It seems likely that IgG1 might have a protective role in clearance of virus from the central nervous system.

To assess the risk of hepatitis B infection among dental personnel, serum samples were collected from dentists of Pune and students, staff, auxiliary staff and class D staff of a dental college in Bombay. Dentists (32.02%), dental auxiliary staff (35.89%), clinical assistants and post-graduate students (19.56%) were found to have significantly higher prevalence of HBV infection as compared to undergraduate dental students (3.94%). The prevalence of HBV infection was high among the dentists as compared to voluntary donors. A positive linear association was observed in the positivity of HBV seromarkers with increasing age and number of years spent by the workers in the dental environment. The rate of increase in HBV seropositivity with age was higher (P less than 0.05) among dental personnel when compared to voluntary donors. Vaccination against hepatitis B is recommended for all the dental students before they start their clinical phase and for susceptible dentists and dental auxillary staff.

Outbreaks of dengue (DEN) fever are reported from 22 villages in five districts in the state of Maharashtra. DEN viral antigen was detected by indirect immunofluorescence test in 34 of 375 female Aedes aegypti mosquitoes collected from 8 villages. DEN types 2 and 3 were identified among the 16 strains that were isolated. None of the 64 male mosquitoes showed the presence of viral antigen; 281 males and 323 females reared from field collected larvae were also found negative. Indoor spraying of DDT in the affected villages produced a drastic reduction in Ae. aegypti catches. No viral antigen was detected in mosquitoes caught subsequent to the spray. With the introduction of water supply through taps, which is often inadequate, people are compelled to store water for domestic purposes, which favours an increase in Ae. aegypti breeding. In view of changing rural ecology associated with the development process, continued surveillance of the spread of Ae. aegypti as well as the diseases borne by this vector are recommended.

Anti-idiotypic antibodies (anti-Ids, Ab2s) were prepared by immunizing rabbits with two murine monoclonal antibodies (Ab1) having specificities for two independent haemagglutinin (HA) epitopes on JE virus [viz., Hs-1, monoclonal antibody (MAb) specific for Japanese encephalitis virus (JEV) and Hx-1, MAb common to flaviviruses]. Anti-Hs-1 (S-Ab2) and Anti-Hx-1 (X-Ab2) reacted specifically with the immunizing Ab1. In addition, they could react with other MAbs whose reactivity was similar to their immunizing homologous Ab1. The paratope inhibition assay indicated that both anti-idiotypes recognized paratope related idiotopes on their respective Ab1 and could therefore be designated as Ab2 beta. Experimental animals (Swiss mice, Balb/c mice and guineapigs) immunized with S-Ab2 or X-Ab2 produced anti-JE virus antibodies (Ab3) which could be detected by enzyme linked immunosorbent assay, immunofluorescence, haemagglutination inhibition and neutralization tests. The anti-idiotypes were also found to stimulate a cellular immune response in vitro as assessed by 3H thymidine incorporation by lymphocytes from JE vaccinated individuals and experimentally immunized Balb/c mice. The findings of the present study suggest that both the anti-Id antibodies are homobodies which may act as surrogate antigens to manipulate the immune response against JEV.

The components of the circulating immune complexes (CICs) were characterised in asymptomatic HIV seropositive individuals. Forty four of 214 individuals (20.56%) showed the presence of CICs. Specific HIV anti-HIV CICs were detected in 33 of them (75%). The isotypic specificity of antibodies found to these CICs was measured. IgG and IgA immunoglobulin classes were detected in CICs.

Sera from healthy donors and four groups of subjects with acute viral hepatitis were tested for anti-HAV IgM by ELISA with hepatitis A virus antigen grown in tissue culture. The results were compared with those obtained by formalin inactivated HAV from 'HAVAB-M' test kit (Abbott Laboratories, USA). With both preparations of antigens, sera from healthy donors and patients suffering from acute hepatitis B or non-A, non-B did not show anti-HAV IgM antibodies whereas acute phase sera from hepatitis A patients showed strong reactions indicating the presence of anti-HAV IgM antibodies. The results indicate similarity in the specificity of both preparations of HAV against anti-HAV antibodies and encourage the use of tissue culture derived HAV for serological diagnosis of hepatitis A.

Outbreak of dengue fever in Chikalthana, Pimpalgaon and Waloor villages in Parbhani district of Maharashtra (India) were investigated. Clinically, the illness was typical of dengue fever except for the absence of maculopapular rash. A total of 42 acute, 14 late acute, 73 convalescent and 19 sera from contacts were collected. Of the 15 virus isolates, 12 were identified as dengue virus type 2 and 1 as dengue virus type 1. Serological tests confirmed the etiological role of dengue virus in the outbreak. House-to-house survey was carried out in Chikalthana and Pimpalgaon villages. Overall, 15.09 per cent of the surveyed population was affected during the outbreak and attack rate was higher at Pimpalgaon. A tendency of water storage was observed in the households and concomitant entomological studies proved Aedes aegypti breeding. Higher prevalence of dengue fever was noted among larger families and in families that had two or more patients, the commonest duration between the first and the last patient was often less than 5 days.

6-MFA, an extract from the fungus Aspergillus ochraceus was administered to 8 bonnet macaques. An equal number of monkeys matched for age, sex and weight received placebo and served as controls. Twenty hours after the administration of the 6-MFA/placebo the monkeys were challenged with an Indian strain of Japanese encephalitis virus by the intranasal route. Signs and symptoms of the disease such as fever, tremors, loss of appetite, dehydration, flaccid paraplegia or quadriplegia were pronounced in all the control monkeys, while in the 6-MFA treated group only two developed symptoms. Virus could be isolated from only one of the 6-MFA treated monkeys on day 6, and from four control monkeys; one each from CSF, spinal cord, blood and from both nasal swab and blood of the fourth monkey. The appearance of HI and N antibodies in 6-MFA treated group was either delayed or completely suppressed. The results indicate that 6-MFA is a potential antiviral agent which can be used to reduce the morbidity and mortality in bonnet macaques (Macaca radiata) experimentally infected with Japanese encephalitis virus.

DNA complementary to the single stranded RNA genome of Chikungunya (CHIK) virus with poly A tract was cloned into the plasmid pGEM-3Zf(-) and 5Zf(+) by blunt end ligation strategy. Clones containing the cDNA inserts were selected by X-gal, IPTG system. They were tested for the expression of structural protein(s) of CHIK virus by in situ enzyme immunoassay and Western blot. The former assay system showed the presence of expressed viral proteins. Analysis of Western blot shows that three structural proteins, E1, E2 and capsid (C) are expressed in Esch. coli. The molecular weights of envelope proteins E1 and E2 were 44-46 Kd and 42-44 Kd respectively, which are lesser than the actual molecular weights of virional proteins (50-52 Kd). This may be due to the absence of glycosylation of these proteins in Esch. coli. In clone no. 382, a high molecular weight protein (56-58 Kd) was observed, which was probably the unglycosylated form of P62 polyprotein coded by the virus during its multiplication. A small protein of MW 6-8 Kd was also expressed in clone nos. 382 and 504, and this appeared to be the unglycosylated form of E3 protein of CHIK virus.

A study of the total and isoenzymes fractions of lactic dehydrogenase (LDH) enzyme was carried out in the plasma of mice infected with Japanese encephalitis (JE) and West Nile (WN) viruses. Only 2 isoenzymes (LDH 4 and LDH 5) were present in control mice. There was a rapid increase of total LDH in the animals infected with the wild strain of viruses. Mice infected with wild strains of JE showed all 5 isoenzymes. LDH 1 disappeared after 12 days of infection but LDH 2 and LDH 3 persisted for 3 wk. The attenuated strain of JE did not produce any change either of the total content of the enzyme or of the isoenzyme pattern. In the plasma of mice infected with wild strains of WN viruses only 4 isoenzyme bands (LDH 2, 3, 4, 5) were detected. The LDH 3 persisted longer than LDH 2. With one strain of WN virus an extra band near LDH 4 was seen.

An Indian strain of Japanese encephalitis virus (JEV), 733913, a human isolate from Bankura, West Bengal in 1973, with all the functional epitopes designated by a panel of murine monoclonal antibodies (MAbs), was treated with one of the JEV specific HI reactive MAb(Hs-I). This led to selection of a neutralization-escape variant which showed loss of reaction to three different MAbs belonging to the same domain (Hs) and assumed similar characteristics to another JEV strain (755468) also isolated from Bankura in 1975 from mosquitoes. It is possible that selection of such variant might occur in presence of pre-existing JE antibody (Hs-I type) in pigs which are amplifying hosts of JEV. Subsequent dissemination of such variant virus could occur through mosquitoes.

A strain of Japanese encephalitis (JE) virus was passaged serially through primary chick kidney cell cultures (45 times) and primary baby hamster kidney cell cultures (21 times). The resultant virus lost its lethal effect to 3 wk old mice by the ic route and 10 day old mice by the ip route. The oligonucleotide fingerprint analysis of the parent and the passaged strains showed 64 spots in common; 17 spots were present in the parent strain which were absent in the passaged virus, while the latter had acquired 9 spots which were not present in the parent virus.