Standardization of serum free media for cryopreservation of RK13 cells.

Foetal calf serum present in the media used for cryopreservation was replaced by various synthetic polymer such as gelatin, glycerol, carboxymethyl cellulose and dimethyl sulphoxide at various concentration. Growth pattern of cells, % survival and karyological studies have been done in the present study. It was found that optimum concentration of carboxymethyl cellulose was 0.1% in combination with 10% glycerol and 10% DMSO. At this concentration percentage survival of cells was found maximum and karyotype was found normal without any abnormality in the chromosomes. It was concluded from the study that serum free media can be employed for the cryopreservation of these cells which are further used for production of tissue culture vaccines without causing any adverse affects.

Epidemiological characteristics of rabies in Delhi and surrounding areas, 1998.

OBJECTIVE: To describe the epidemiological characteristics of rabies in Delhi in 1998. METHODS: Analysis of the records of hydrophobia cases admitted to the Infectious Diseases Hospital, Delhi (IDH) in 1998. RESULTS: About 46 percent (99/215) of the hydrophobia cases admitted to the IDH in 1998 belonged to Delhi. The remaining came from the adjoining states, both urban and rural areas. In Delhi residents, overall hospitalization rate was 0.81 per 100,000 population. It was significantly higher in 5-14 year old than in other age groups and in males than in females (p <0.0009). Cases occurred round the year. Almost 96 percent cases (206/215) gave history of animal exposure, 13 days to 10 years (median 60 days) before hospitalization. Majority (195/206) had class III exposure. Animals involved were stray dog (193/206 = 90 percent), pet dog, cat, jackal, mongoose, monkey and fox. Most of cases were never vaccinated (78 percent) or inadequately vaccinated (22 percent); only 1 percent each received appropriate wound treatment, or rabies immunoglobulin. CONCLUSIONS: Rabies is a major public health problem in Delhi. Its incidence is significantly higher in 5-14 year old children than in other age groups. The results indicate the need to educate the community and health care workers about the importance of immediate and adequate post-exposure treatment and to start an effective control program for dogs, the principal vector of rabies.

Moderate immunization coverage levels in East Delhi: implications for disease control programmes and introduction of new vaccines.

A vaccination coverage survey conducted in East Delhi in September 1999 showed that only 58.6 per cent of the children aged 12-23 months had received the full course of the vaccines recommended under the national immunization programme. Coverage with the third dose of DTP and oral polio vaccines was around 71 per cent, and with BCG and measles vaccines was 83 and 59 per cent, respectively. Drop-out rates between DTP1 and DTP3 and between DTP1 and measles immunization were 13.8 and 28.7 per cent, respectively. Nine per cent of the children had not received a single dose of any vaccine. The main reason for failure to immunize was lack of information. There was a marginal increase in DTP3 and OPV3 immunization coverage levels as recorded through a previous survey in 1996, a drop in coverage with measles vaccine from 64.3 to 59 per cent, and a significant increase in tetanus toxoid immunization coverage of pregnant women from 79.4 to 93 per cent. The percentage of children who had not received any vaccine declined from 13 to 9 per cent in the period between the two surveys. Coverage with hepatitis B vaccine at 14 per cent was only marginally higher than the baseline rate of 9 per cent before the vaccine was made available, free of cost, through government and municipal corporation health facilities.

Diethylene glycol poisoning in Gurgaon, India, 1998.

OBJECTIVE: To discover the cause of acute renal failure in 36 children aged 2 months to 6 years who were admitted to two hospitals in Delhi between 1 April and 9 June 1998. METHODS: Data were collected from hospital records, parents and doctors of the patients, and district health officials. Further information was obtained from house visits and community surveys; blood and stool samples were collected from other ill children, healthy family members and community contacts. Samples of drinking-water and water from a tube-well were tested for coliform organisms. FINDINGS: Most of the children (26/36) were from the Gurgaon district in Haryana or had visited Gurgaon town for treatment of a minor illness. Acute renal failure developed after an episode of acute febrile illness with or without watery diarrhoea or mild respiratory symptoms for which the children had been treated with unknown medicines by private medical practitioners. On admission to hospital the children were not dehydrated. Median blood urea concentration was 150 mg/dl (range 79-311 mg/dl) and median serum creatinine concentration was 5.6 mg/dl (range 2.6-10.8 mg/dl). Kidney biopsy showed acute tubular necrosis. Thirty-three children were known to have died despite being treated with peritoneal dialysis and supportive therapy. CONCLUSION: Cough expectorant manufactured by a company in Gurgaon was found to be contaminated with diethylene glycol (17.5% v/v), but a sample of acetaminophen manufactured by the same company tested negative for contamination when gas-liquid chromatography was used. Thus, poisoning with diethylene glycol seems to be the cause of acute renal failure in these children.

Community studies on hepatitis B in Rajahmundry town of Andhra Pradesh, India, 1997-8: unnecessary therapeutic injections are a major risk factor.

In Rajahmundry town in India, 234 community cases of jaundice were interviewed for risk factors of viral hepatitis B and tested for markers of hepatitis A-E. About 41% and 1.7% of them were positive for anti-HBc and anti-HCV respectively. Of 83 cases who were tested within 3 months of onset of jaundice, 5 (6%), 11 (13.3%), 1 (1.2%), 5 (6%) and 16 (19.3%) were found to have acute viral hepatitis A-E, respectively. The aetiology of the remaining 60% (50/83) of cases of jaundice could not be established. Thirty-one percent (26/83) were already positive for anti-HBc before they developed jaundice. History of therapeutic injections before the onset of jaundice was significantly higher in cases of hepatitis B (P = 0.01) or B-D (P = 0.04) than in cases of hepatitis A and E together. Other potential risk factors of hepatitis B transmission were equally prevalent in two groups. Subsequent studies showed that the majority of injections given were unnecessary (74%, 95% CI 66-82%) and were administered by both qualified and unqualified doctors.

Silent spread of dengue and dengue haemorrhagic fever to Coimbatore and Erode districts in Tamil Nadu, India, 1998: need for effective surveillance to monitor and control the disease.

Dengue fever (DF) or dengue haemorrhagic fever (DHF) has not previously been reported in Coimbatore and Erode districts in Tamil Nadu in India. In 1998, 20 hospitalized cases of fever tested positive for dengue virus IgM and/or IgG antibodies. All of them had dengue-compatible illness, and at least four had DHF. Two of them died. Sixteen cases were below 10 years of age. The cases were scattered in 15 distantly located villages and 5 urban localities that had a high Aedes aegypti population. Although the incidence of dengue-like illness has not increased recently, almost 89% (95/107) of samples from healthy persons in the community tested positive for dengue IgG antibodies. The study showed that dengue has been endemic in the area, but was not suspected earlier. A strong laboratory-based surveillance system is essential to monitor and control DF/DHF.

Vaccines: quality issues.

PIP: Globally, vaccines are administered to a number of healthy individuals, mostly infants, in the National Immunization Programs (NIPs). Hence safety and quality issues are of paramount importance. Problems in vaccine quality can have a major negative impact on NIPs. Unsafe or ineffective vaccines threaten to disable vaccine delivery systems that might otherwise prevent the deaths of millions of infants and children each year. This article explores quality issues related to vaccines. The paper begins with a brief review of how poliomyelitis has been successfully eradicated globally. It then discusses vaccine production, control and regulation issues. The functions of the National Control Authority/National Regulatory Authority (NRA) with regard to vaccine production are outlined. These control functions are designed to ensure safety and efficacy of vaccines produced and to guarantee the consistency of production: 1) a published set of requirements for licensing, 2) surveillance of vaccine field performance, 3) system of lot release, 4) use of the laboratory when needed, 5) regular infections for good manufacturing and laboratory practices, and 6) evaluation of regular performance. Finally, the paper illustrates how locally blended oral polio vaccine and vaccine vial monitors has been successfully implemented in India and the role of NRA in its success. It is noted that the vaccine used in NIP is safe and efficacious as the number of reported cases of polio has declined dramatically in 1998 and 1999.^ieng

A survey on community perceptions of jaundice in east Delhi: implications for the prevention and control of viral hepatitis.

Using senior health professionals as interviewers, a 30-cluster sampling survey was carried out to investigate community perceptions of pilia (the local word for jaundice) in east Delhi (India). Of 416 persons (mostly mothers of children aged < 2 years) interviewed, 339 (81%) were aware of pilia as an illness. Only 322 (77%), 164 (39%), 73 (18%) and 71 (17%) people knew about correct symptoms, dangers, causes and prevention of pilia. Most of the correct responses were related to the faeco-orally transmitted viral hepatitis. Literate respondents were significantly more aware of pilia (chi 2 52.81, P < 0.0001), its symptoms (chi 2 48.88, P < 0.0001), causes (chi 2 39.34, P < 0.0001), dangers (chi 2 19.3, P = 0.0007), and prevention (chi 2 60.8, P < 0.0001). However, age of the respondents had no significant bearing (P > or = 0.05) on the correctness of responses. About 293 (70%) subjects considered pilia as a treatable illness; of them, 193 (66%) and 77 (26%) respectively expressed their preference for the 'modern' and indigenous systems of medicine for its treatment. In contrast, 110 (38%) respondents said that they would prefer faith healers for the treatment of pilia. Although only 31 (7%) persons were aware of a vaccine against pilia (hepatitis B vaccine), virtually all agreed to have their children immunized if such a vaccine were made available. The study underscores the usefulness of pilia in lay-reporting of viral hepatitis and epidemiological studies on jaundice-associated illnesses and the need for educating the community about its causes and prevention to increase people's participation in controlling viral hepatitis and other diseases that mainly manifest as jaundice.

Community studies on prevalence of HBsAg in two urban populations of southern India.

OBJECTIVE: To find out prevalence of HBsAg in general population, especially in under-five children. SETTING: Bangalore and Rajahmundry towns in southern India. METHODS: Localities were chosen as the sampling units in each town. About 10-20 households were randomly selected from each locality. Only the youngest but apparently healthy person present in the household was interviewed for age, sex and history of jaundice any time in life. Mothers were interviewed to collect data for children below 15 years of age. Blood samples were collected from these persons on filter paper strips (18-mm diameter disc, Whatman filter paper No. 3) by finger prick method. The samples were tested for HBsAg by Micro ELISA (Ortho-Clinical Diagnostics). RESULTS: Overall, 3.3% (95% CI, 2.0-4.5) of 737 persons in Rajahmundry and 4.2% (95% CI, 2.8-5.5) of 816 persons in Bangalore were found carriers of HBsAg. Age-specific or sex specific carrier rates were similar in Rajahmundry as well as in Bangalore. Most of the carriers (96%) denied having jaundice ever in life. CONCLUSIONS: The results from this community based study are in agreement with the historical data from hospital based studies that about 3-5% of persons may be carriers of HBsAg and that the pool of chronic carriers of hepatitis B virus in India is built up in childhood and is then maintained in older children and adults. The results highlight the need of completing hepatitis B immunization during the infancy.

Insecticide susceptibility status of Culex tritaeniorhynchus giles, vector of Japanese encephalitis in Delhi.

Laboratory studies were carried out to ascertain the current susceptibility status of adult and larval stages of the Culex tritaeniorhynchus mosquito, vector of Japanese encephalitis, to various insecticides used under public health programs in India. The present study revealed that exposure of adult mosquitoes to diagnostic concentrations of DDT - 4.0%, malathion - 5.0%, fenitrothion - 1.0%, and propoxur - 0.1% could induce only 50.0, 10. 0, 15.0, and 5.0% mortality, respectively, indicating that the species was resistant to all of these insecticides. The LT50 and LT95 values calculated using diagnostic concentrations of DDT, malathion, fenitrothion, and propoxur were found to be 56.4 and 136, 138 and 272, 185 and 258, and 187 and 249 min, respectively. However, when adult mosquitoes were exposed to the diagnostic concentration of synthetic pyrethroids, viz., deltamethrin - 0.025%, permethrin - 0.25%, and lambdacyhalothrin - 0.1%, 100.0% mortality was observed, indicating that the species was highly susceptible to these adulticides. Larval susceptibility tests carried out using diagnostic dosages of DDT- 0.008, temephos- 0.02, fenthion- 0.008, fenitrothion- 0.125, and malathion- 0.005 mg/l failed to induce any mortality, indicating that larvae were resistant to these larvicides. The LC50 and LC90 values calculated for commonly used larvicides, viz., temephos and fenthion, were 0.1511 and 1.9098, and 0.6151 and 2.395 mg/l, respectively. Increase in tolerance level were estimated at 95.5- and 299.4-fold when these LC90 values were compared with diagnostic dosages of temephos and fenthion, respectively.

A severe and explosive outbreak of hepatitis B in a rural population in Sirsa district, Haryana, India: unnecessary therapeutic injections were a major risk factor.

Most outbreaks of viral hepatitis in India are caused by hepatitis E. This report describes an outbreak of hepatitis B in a rural population in Haryana state in 1997. At least 54 cases of jaundice occurred in Dhottar village (population 3096) during a period of 8 months; 18 (33.3%) of them died. Virtually all fatal cases were adults and tested positive for HBsAg (other markers not done). About 88% (21/24) of surviving cases had acute or persistent HBV/HCV infections; 54% (13/24) had acute hepatitis B. Many other villages reported sporadic cases and deaths. Data were pooled from these villages for analysis of risk factors. Acute hepatitis B cases had received injections before illness more frequently (11/19) than those found negative for acute or persistent HBV/HCV infections (3/17) (P = 0.01). Although a few cases had other risk factors, these were equally prevalent in two groups. The results linked the outbreak to the use of unnecessary therapeutic injections.

Diphtheria is declining but continues to kill many children: analysis of data from a sentinel centre in Delhi, 1997.

Although diphtheria is declining in Delhi, case fatality rates (CFRs) are rising. In 1997, of 143 clinically suspected cases admitted to the Infectious Diseases Hospital 45 (32%) died. We examined their records to understand the epidemiology and reasons for high CFRs. About 53% of cases were from Delhi; they were not limited to any particular area. All the deaths and 92% (131/143) of cases occurred in children below 10 years of age. Only 12% of cases had received one or more doses of DPT. Muslims contributed significantly more cases than Hindus. CFRs were significantly higher in young (P = 0.03) and unvaccinated (P = 0.01) children and in those who received antitoxin on the third day of illness or later (P = 0.03). The study highlights the importance of improved vaccine coverage and early diagnosis and prompt administration of antitoxin in reducing CFRs for diphtheria in Delhi.

Epidemiological characteristics of poliomyelitis in Delhi, 1997.

OBJECTIVE: To describe the epidemiological characteristics of poliomyelitis in Delhi in 1997 after four consecutive statewide immunization campaigns with oral polio vaccine (OPV). METHODS: Stool samples were collected from 158 cases of acute flaccid paralysis (AFP) along with their age, sex, residential address, immunization history and dates of onset of paralysis, reporting and investigation. The samples were processed for isolation of polioviruses. In addition, historical data on vaccination coverage surveys and OPV testing were reviewed. These data were analyzed to understand the epidemiological patterns of poliomyelitis in Delhi. RESULTS: Of 158 cases of AFP, about 23% were investigated within 2 days of onset of paralysis. Two samples each were collected from 97 (61%) cases, and one each from the remaining cases. Detection of 158 cases of AFP gave an incidence of 1.34 per 100,000 population. About 36% (57/158) of AFP cases excreted poliovirus, mostly (53/158) wild poliovirus. Of the wild poliovirus isolates, 72% (38/53) and 25% (13/53) were serotypes P1 and P3 respectively; 2 isolates were P2. Almost 95% (146/154) of AFP cases and all the laboratory confirmed cases (excreting wild poliovirus) occurred in children below 5 years of age. Only one-third of AFP (55/158) or laboratory confirmed cases (18/53) had received 3 or more doses of OPV before onset of paralysis. About one-fourth of cases in both the categories were totally unvaccinated. AFP cases occurred round the year but peaked in November-December. Peaks were always observed during July-August in the past. The cases were widely scattered without any obvious clustering in any locality. CONCLUSIONS: Poliomyelitis has declined substantially in Delhi. The study underscores the need for further efforts to improve vaccine coverage levels, AFP surveillance, and cold chain maintenance to achieve the complete interruption of transmission.

Alternative approach and evaluation of three potency assay methods for testing adsorbed tetanus toxoid.

Thirteen batches of adsorbed Tetanus Toxoid (TT) from different manufacturers were tested for potency by three different methods viz: (i) An Antibody Induction Method (AIM) developed in mice: (ii) WHO lethal challenge in mice; and (iii) Conventional Antibody Induction (I.P). Method in guinea pigs. The potency results obtained in AIM, by serological evaluation of immunized mice were found identical and correlated significantly with those obtained by WHO recommended lethal challenge test in mice. The potency data obtained in the present study was found comparable with other studies. An AIM in mice thus offers an alternative to lethal challenge tests and can replace guinea pig model. Out of 107 serum samples obtained from immunized guinea pigs in the conventional antibody induction method, 90% samples contained more than 4 units of tetanus antitoxin per ml. End point titres of 42 serum samples belonging to 5 batches of TT also showed much higher tetanus antitoxin content when determined by TN test. The potency data obtained thus suggest revision of the minimum requirement in Indian Pharmacopoeia which is too low and which may be increased as indicated by the present study.

Quantitation of tetanus and diphtheria antitoxins in mouse sera by indirect haemagglutination.

Serum samples obtained from 75 groups of mice immunized with various doses of adsorbed tetanus vaccine, adsorbed diphtheria-tetanus vaccine and adsorbed diphtheria-tetanus-pertussis vaccine were titrated for tetanus antitoxin content by an in-vitro indirect haemagglutination (IHA) and by toxin neutralization test (TN) in mice. From these serum samples of 49 groups of mice which were immunized with combined vaccine containing diphtheria toxoid were titrated for their diphtheria antitoxin content by IHA and by i.d. toxin neutralization test (TN) in guinea pigs. Good correlations were found between the estimates obtained by in-vitro IHA and in vivo TN tests in both tetanus and diphtheria antitoxin titrations. The minimum level of tetanus or diphtheria antitoxin detectable by IHA was 0.00039 IU/ml. It is concluded that IHA is a simple, sensitive and reproducible alternative test which can replace the animal TN tests for the estimation of tetanus and diphtheria antitoxins and could reliably be used in the potency assay of tetanus and diphtheria toxoids of combined vaccines based on antibody induction in mice.

Endemic cholera in Delhi, 1995: analysis of data from a sentinel centre.

Data on cholera cases admitted to the Delhi Infectious Diseases Hospital (IDH) are presented to describe the pattern of occurrence of cholera in Delhi in 1995. Rectal swabs from 4082 cases of acute diarrhoea admitted to the IDH were examined for excretion of Vibrio cholerae. Of them, 2004(49%) and 4(0.1%) were positive for V. cholerae O1 biotype El Tor and V. cholerae O139 respectively. Most cholera cases occurred during May-September (summer and monsoon months). The period from January to March (winter) was completely free from cholera. The urban areas were not affected uniformly. Of the 80 PIN (Postal Index Number) code areas, 10 contributed to 57% of the cases. The early cases were scattered in PIN code areas distant from one another. The hospitalisation rates for cholera were the highest in children aged less than five years and declined significantly with increasing patients' age. Males had significantly higher rates than females aged up to 20 years, whereas the situation was reversed in the 20 to 39 year age group. Four per cent of the affected families had multiple cases. An estimated 1% of the household contacts of hospitalised cases of cholera were themselves hospitalised for cholera within 2 days of the first admission. Of the 260 V. cholerae O1 isolates tested, 4%, 7%, 8%, 89%, 91% and 95% were resistant to tetracycline, nalidixic acid, chloramphenicol, co-trimoxazole, streptomycin, and furazolidone respectively. The study highlights the usefulness of surveillance data to identify groups, urban areas and seasons with increased risk for cholera and to allow control measures to be focussed on those in greatest need.

Outbreak of viral hepatitis B in a rural community in India linked to inadequately sterilized needles and syringes.

In India, virtually all outbreaks of viral hepatitis are considered to be due to faeco-orally transmitted hepatitis E virus. Recently, a cluster of 15 cases of viral hepatitis B was found in three villages in Gujarat State. The cases were epidemiologically linked to the use of inadequately sterilized needles and syringes by a local unqualified medical practitioner. The outbreak evolved slowly over a period of 3 months and was marked by a high case fatality rate (46.7%), probably because of concurrent infection with hepatitis D virus (HDV) or sexually transmitted infections. But for the many fatalities within 2-3 weeks of the onset of illness, the outbreak would have gone unnoticed. The findings emphasize the importance of inadequately sterilized needles and syringes in the transmission of viral hepatitis B in India, the need to strengthen the routine surveillance system, and to organize an education campaign targeting all health care workers including private practitioners, especially those working in rural areas, as well as the public at large, to take all possible measures to prevent this often fatal infection.

PIP: Viral hepatitis outbreaks in India are generally due to feco-orally transmitted hepatitis E virus. Described in this paper is a cluster of 15 cases of viral hepatitis B found in three villages in Gujarat State's Mehasana District linked to the use of inadequately sterilized needles and syringes by an unqualified medical practitioner. Patients were identified by house-to-house surveys and blood tests. The outbreak, which evolved slowly over a 3-month period, had a 46.7% case fatality rate because of concurrent infection with hepatitis D infection and sexually transmitted diseases. Without fatalities, this outbreak would not have been noticed. This event indicates a need to strengthen the routine hepatitis B surveillance system in India as well as to target health care workers in rural areas for an educational campaign about the importance of sterile equipment.

Epidemiologic consequences of moderate coverage levels of measles vaccine in a district headquarter town (Alwar) in India, 1996.

This paper describes the epidemiology of measles in a medium size town (population 240,000) in India where vaccine coverage levels have remained constant at around 70 per cent in the past 7 years. A retrospective community survey covering 4023 children under 10 years old detected 252 cases of measles in the previous year. This gave an annual incidence of 6.3 per cent (95 per cent CI 5.5-7). About half of the cases occurred in vaccinated children. Only 5 per cent of the cases occurred in children below 9 months of age. This age is appropriate for routine measles immunization. Despite modest coverage levels with only 54 per cent effective vaccine (estimated by a screening method), there was a modest upward shift in the age distribution of measles cases; the median age was more than 48 months.

Merit of mobilization technique in transfer of non-autotransferable R-plasmids.

A total of 200 urinary isolates of Esch.coli received at National Salmonella and Escherichia Centre, Central Research Institute, Kasauli during the years 1995 to 1997 were studied for transferable drug resistance. Out of 188 strains, 134 strains showing resistance to either Nalidixic acid or Rifampicin were subjected to autotransferable resistance studies. Of these 134 strains 131 showed either partial or enbloc transfer of R-factor. Mobilization experiment successfully transferred resistance marker in 14 of the 68 isolates in which resistance to one or more drugs could not be transferred during conjugation experiment.