Examination of only one sputum sample cannot detect all the sputum positive cases in the community. To obtain better estimates of the prevalence of bacteriologically confirmed disease in the community, a study was conducted to find out the additional yield of cases through collection and examination of eight sputum specimens and also in order to work out correction factors for estimates based on one or two sputum samples, as collecting multiple sputa is very difficult. The study was carried out in 77 villages in Nelamangala Taluk of Bangalore. In all, 5826 persons were referred for sputum examinations.

Results of all the eight culture examinations were available for 2973 (51% of the eligibles). Of these 64 persons were positive by culture of atleast one specimen. Each of the eight specimens has the chance of detecting a case and any one of them could be considered as first or second specimen etc. To overcome this difficulty 80 permutations were randomly chosen out of the total 40,320 permutations possible. Cases from first specimen and additional cases from subsequent specimens were calculated through four mathematical equations. The first equation namely Y = KXm (28.66 x-1.40) has been considered as providing the best fit to the observed data. On the basis of this equation it appears that additional positives could be obtained upto the 1Oth specimen. Out of 64 culture positive cases, only 72% of positives could be detected by first two samples. To get about 95% of the cases, it is necessary to examine at least six specimens from each individual. Multiple samples are rewarding for detecting even high grade cultures.

This paper reports on a study conducted in the year 1975 to estimate yield of tuberculosis cases from multiple sputum specimens, and work out correction factors to be applied to estimates based on small number of specimens. Eight sputum specimens were collected within a fortnight from each person with an abnormal chest X-ray during an epidemiological survey in 77 villages in a district of south India. Each specimen was examined by Ziehl-Neelsen technique of microscopy and culture. In all, 3,199 persons were referred for sputum examination and results of all the eight specimens were available for 1,652. Of the latter, 64 were culture positive.

The first specimen detected 58% of the culture positives and the additional positives by later specimens generally decreased. The contribution from the first specimen was 71% for cultures showing good growth and 19% for cultures with scanty growth. Similarly for positives on both culture and microscopy, first specimen detected 87% whereas the corresponding proportion was 32% for those positive only on culture. The type of specimen (viz., spot or overnight) and age or sex of the case did not influence the yield from multiple examinations. The precision of an estimate of prevalence will depend on the number of specimens on which it is based and the coverage obtained in the collection and examination of specimens. Correction factors to be applied to such estimates based on one or two specimens, for various levels of coverage have been presented. For example, an estimate of prevalence based on one sputum specimen with 90% coverage will have to be nearly doubled to get a more precise estimate. Using these correction factors, revised estimates of prevalence have been presented for a number of prevalence surveys conducted in India. It has been estimated that the total number of infectious cases in India at present may be at least 3 million, as against 2 million according to earlier estimates.

A study was conducted in a slum area of Bangalore, to get information on the prevalence of all forms of tuberculosis in 0-4 year age group, respiratory tuberculosis in 5-14 year age group and the proportion of respiratory tuberculosis among total respiratory diseases in 0-14 year age group. Entire population in a slum area was investigated. Children aged 0-9 years were given tuberculin test and their nutritional status assessed. All persons were X-rayed. Sputum specimens were collected from those having radiological abnormality in chest, chest symptoms of one week or more in 0-4 years, in addition from those with any kind of sickness, malnutrition and tuberculin reactors.

In 0-9 year age group, 5.5% were tuberculin positive (without BCG lesions), in 0-4 years, 1.8% and 5-9 years, 11.3%. Among the X-rayed children, 47.4% had some kind of sickness, the proportion being significantly high in 0-4 year age group. The respiratory sickness is the commonest among children of all ages followed by malnutrition (21%). Among children with chest symptoms, upper respiratory infections were 33%. Chest X-ray abnormalities were present in 4.5% of children and of these 82.5% had non-specific pneumonitis. Of 71 persons with respiratory disease, about 7% were tuberculous. Out of 1408 children, only 5 had active primary tuberculosis, giving a prevalence of 0.35%. None in 0-4 year age had sputum positive disease or extra pulmonary tuberculosis.

It has been highlighted that non-tuberculous chest diseases are common in pediatric age group and many of these may be wrongly classified as active tuberculous in practice. It is concluded that tuberculosis in the pediatric age group in this community is not a serious public health problem.

The findings of the two follow-ups conducted passively, in a slum area of Bangalore city, each at an interval of one year, are presented here. The objectives of the follow-ups were to study the migration, episodes of sickness and health status of sick people of initial survey over a period of time in the pediatric age group. The information was collected by two ways: (1) by visiting each house to collect information on sickness among them during the preceding year as per the questionnaire. (2) Going through the records of the Area Health Centre about various morbidities among the residents of the area pertaining to symptoms, diagnosis and treatment during any year noted.

Migration had occurred upto 7% in 0-9 year age group within first year and no migration during second year in 0-4 year age group. Out of 400 children belonging to 0-14 years, had symptoms related to respiratory system. In all, 5 children were diagnosed as case of active primary tuberculosis, non-e died in two years and one had persistent respiratory symptoms. Hospital records showed that only 1 out of 5 had attended any health facility with respiratory symptoms. The usefulness of passive follow-up without clinical investigations as a tool, needs to be reviewed. Most of the symptomatics do not go to hospital. Diagnosis cannot be arrived at by passive follow-up. It may be necessary to decrease the interval of follow-ups if it is desired to get precise idea on frequency of episodes of sickness, as most of the sicknesses are forgotten by the population with passage of time.

The entire population of a slum area of Bangalore city, comprising of 3313 persons was registered, questioned for symptoms and offered chest X-ray at a centre located in the slum itself. Those, who had any chest symptom and/or X-ray abnormality, were offered detailed examinations, viz., clinical examinations, repeated examinations of sputum for tubercle bacilli, and further chest X-rays. Of the total 2855 persons X-rayed and/or questioned, 1039 needed detailed examinations and about a fifth of the latter required referral to a consultant panel for diagnosis of chest diseases. Further, about 60% of those referred to consultants needed special investigations. Thus, the study of prevalence of chest diseases in the community needed considerable facilities and were operationally difficult. It is envisaged that similar problems will also be faced if peripheral dispensaries are to make proper diagnosis of chest diseases, due to the need for referral of large number of patients and provision of complicated diagnostic facilities at the referral hospitals. The study seeks to quantify the problem of chest diseases and tuberculosis in the slum community.

The prevalence of sickness in the population at any point of time were 49.5%. Sickness related to the respiratory system was 13.3%. It increased with age and was highest (42.6%) in those aged 55 years and above. Among 2855 persons X-rayed, 145(5.1%) had any radiological abnormality in chest. It is seen that respiratory systems symptoms were commonest in all the age groups. A total of 172 patients were diagnosed to have respiratory system abnormalities with or without X-ray lesions. Of them, 75% had non- tuberculous etiology, 7.6% had active pulmonary tuberculosis and the remaining 17.4% had inactive tuberculosis. Prevalence of sputum positive cases was 0.26% and prevalence of total active pulmonary tuberculosis was 0.44%. The problem of arriving at final diagnosis was dependent on application of complicated special investigation tools to a large community. In view of the low coverage (47.4%) for the special investigations, prevalence of different chest diseases in the community could not be investigated.

It is concluded that in the community under study, the size of the problem of non- tuberculous diseases of the chest and operational problems in their diagnosis were considerable.

Prevalence of tuberculous infection in young children is an important surveillance measure. However, the hypersensitivity may be depressed by malnutrition and thus interfere with the interpretation of tuberculin test leading to underestimation of the infection rate. Objective of this investigation was to study the relationship between tuberculin reaction with 1 TU RT 23 and nutritional status of children. The study was carried out in 1974 among children aged 1-9 years of age living in an urban slum area of Bangalore city and who were not given BCG vaccination.

Of the 1151 registered children aged 0-9 years, 482 in the age group 1-4 and 526 in 5-9 years formed the study group. Of these 1008 children, 980 had both clinical evaluation and anthropometric measurement for nutritional status and 963 had both tuberculin test readings and anthropometric measurements carried out for them. Of the 482 children aged 1-4 years, 230 were classified as suffering from Protein Calorie Malnutrition (PCM) and of the 498 in the 5-9 years of age, 227 were classified as suffering from PCM. Distribution of tuberculin test indurations in mm among the normals and the undernourished were compared; no significant difference in the mean size of tuberculin indurations as well as in the distributions of these indurations was observed, regardless of the method used for arriving at the classification.

In the District Tuberculosis Programme (DTP) the diagnosis is based on sputum microscopy. Majority of health institutions in the district are provided with microscopes for this purpose. In the Peripheral Health Institutions, the programme activities have to be carried out by its staff after a short period of training given by District TB Centre personnel on the spot. So the microscopy work in the PHIs is likely to be carried out by any paramedical personnel and not necessarily by a qualified laboratory technician. It is therefore, necessary to know whether the standard of microscopy carried out by these paramedical personnel after a short training will be upto the mark. To assess the efficiency of smear examination done by these individuals, a study was conducted in Bangalore district covering nine microscopy centres in various types of health institutions, a few months after the implementation of the programme. Under the DTP a spot specimen is collected from every chest symptomatic attending the health institutions and a smear is made and examined for the presence of AFB and all positive cases are put under treatment. The sputum specimens and the smears examined in these nine centres were brought to National TB Institute laboratory. The smears were examined by an experienced laboratory technician. Duplicate smears were also prepared from these specimens and their results compared with results of re examination and centre's examination. All specimens were cultured by swab method and all positive cultures were subjected to sensitivity and identification tests.

Analysis of the results based on culture showed that barring a few centres where the performance was poor, the standard of examination was fairly good. The under and over diagnosis based on culture were 38.2% and 2.6% respectively, and these were within the limits observed generally. Comparison of results on re examination of centre smears and duplicate smears indicated that both reading variation and defective smear preparations and staining could have influenced under diagnosis in these centres. The study has also thrown some light on methodology of assessment of sputum examination that could be adopted wherever a tuberculosis control programme is functioning.

This paper deals with the cost of establishing and running a bacteriological laboratory in State Tuberculosis Centres under the National Tuberculosis Programme, and the cost of various examinations to be undertaken in such a laboratory. A knowledge of the cost will enable proper planning and judicious utilization of the resources. Further, when services are rendered to private individuals or institutions, the charges for different examinations can be levied on a rational basis. The place of smear and culture examinations under the programme, the implications of establishing a culture laboratory, the limitations of cost worked out, have been discussed. A plan of the laboratory building is also provided.

The cost has been worked out presuming that about 12,000 specimens per year are likely to be received, of which 25% might turn out to be positives. Non recurring cost was estimated to be Rs.1,07,724 and annual recurring cost would be Rs.49,709. Factors that contribute to the cost structure are overheads, cost of material and labour. In addition, certain essential facilities like cold room, incubator room, gas supply, washing and sterilisation etc., add to the cost. (i) staff-bacteriologist-1, lab technicians-4, lab attendants-3 and registration clerk-1; their salaries, (ii) building-rent (iii) electricity (iv) furniture (v) equipment and supplies (vi) water charges had all been taken into consideration. The cost of one smear examination was estimated to be Rs.0.54 and that of culture and sensitivity test Rs.9.43.

A study was conducted to evaluate the usefulness of pyruvate medium for isolation of M.bovis from human material and additional yield of M. tuberculosis resistant to INH. Specimens from both rural and urban populations were included for this study in order to understand the problem in both the situations. There were two studies in progress at the National Tuberculosis Institute when pyruvate media slopes were introduced for culture purpose. One study was an epidemiological survey; 2518 sputum specimens received from 51 villages covering a population of about 32,300 were used. The specimens were collected from persons aged 5 years and above showing abnormal shadow on X-ray. The other study was conducted in collaboration with the State Tuberculosis Centre, Bangalore; 1204 sputum specimens were received from out patients attending the centre. In addition to LJ medium, pyruvate medium was used for isolation purposes. Identification and sensitivity tests were done on positive cultures as per routine. In all, 129 cultures of tubercle bacilli were isolated from 2118 specimens belonging to study 1 and 398 from 1204 specimens belonging to study 2. The number of cultures contaminated were 253 and 35 respectively. No M.bovis was isolated in either study. There were 24 and 23 cultures resistant to INH among those isolated from LJ and pyruvate medium respectively. Thus, no increase was observed in the isolation of INH resistant strains using pyruvate medium.

Hence, no benefit was derived by using this medium over and above what was obtained from plain Lowenstein Jensen medium in both the situations.

Sputum microscopy is the main casefinding tool in tuberculosis control programmes. The technique of smear preparation is an important step which needs to be simple for wide applicability. It is often stressed that smear should be prepared from the purulent portions of the sputum as they are likely to have more number of bacilli. It may not be possible for the microscopist/paramedical worker at the periphery to strictly follow this procedure. Hence, a study was conducted to compare the sensitivity of 4 methods of sputum smear preparation viz., direct smear prepared (i) blindly without making any selection of portions of sputum specimen, (ii) from portions of sputum material likely to contain the bacilli, (iii) after mixing up the sputum specimens thoroughly, and (iv) from centrifuged deposit after homogenization of sputum with sodium hydroxide and concentration by centrifugation. Culture was also done for Mycobacterium tuberculosis.

A total of 549 specimens were employed. Positivity rates by four methods were: 79.6% by method (i), 80.3% by method (ii), 80.7% by method (iii) and 77.2% by method (iv). There was no statistically significant difference in the number of positives obtained from different methods. Centrifuged deposit smears proved to be in no way better than the direct smears. The differences in the methods lay only in the classification of positive smear as of a low or high grade.

Studies from Madras had shown that the strain of M.tuberculosis isolated from south India were low virulent to guineapigs. The relationship between virulence in guineapigs and pathogenesis in humans could not be established earlier. A study was conducted to investigate the relationship of virulence with the pathogenesis by comparing the virulence of isolates from pulmonary tuberculosis with that from patients with TB meningitis. The strains of bacilli were obtained from three different sources: a) Sputum from rural tuberculosis patients living near Bangalore city, b) sputum of TB patients living in the city and c) from Cerebrospinal fluid (CSF) of patients suffering from tuberculous meningitis and admitted in different institutions in Bangalore city. The specimens were processed by standard recommended procedures and cultured on Lowenstein Jensen medium. The identification of an isolate as M. tuberculosis was based on the niacin test. Albino Guinea pigs of both sexes (who were bred and raised at this Institute) were used for the tests. The virulence assay and the calculation of the root-index of the virulence (RIV) were carried out according to the Mitchison method.

1) As per the RIV method, virulence has been classified into low, moderate and high. The study showed that the percentages of cultures having isolates of low, moderate and high virulence, were the same as that of isolates obtained from patients in Madras, reported by Mitchison et al., in 1960. 2) The distribution of the RIV of virulence of isolates from patients living in the city of Bangalore was significantly different (p < 0.05) from that of isolates from patients living in rural Bangalore. 3) The number of cultures classified as high virulent were significantly greater in isolates from patients with tuberculous meningitis compared with those from patients with pulmonary tuberculosis. However, 36% of the isolates from the meningitis group were of low virulence.

A study was done to compare (1) the filtration method with conventional centrifugation method for the recovery of tubercle bacilli from urine and (2) drug sensitivity profile, virulence for guinea pigs and phage type of the urine isolates with the corresponding isolates from the sputum of cases of bacillary pulmonary tuberculosis.

Urine specimens from 236 pulmonary tuberculosis patients were cultured by routine centrifugation method as well as filtration method. Filtration was done by passing urine through a 0.45 um membrane filter and treating the membrane with 5% oxalic acid for 15 minutes. LJ medium was used for culture in both the methods. Centrifugation yielded 27 positives (11.6%) whereas filtration gave 12 (12.6%) out of 95 specimens filtered. Contamination was more with filtration method. Comparison of the biological properties of M.tuberculosis isolated from urine and sputum of the same patients revealed difference in drug sensitivity profile or virulence for guineapigs for 13 of 25 (52%) of the pairs of isolates tested. Moreover 4 of 11 pairs subjected to phage typing were found to differ in both major and minor phage types. The significance of these findings in the light of the pathogenesis of tuberculosis is also discussed.

One of the hypothesis put forth for the failure of BCG vaccine to show protection against bacillary pulmonary tuberculosis in Chingleput trial was the interference from non-tuberculous mycobacteria that were prevalent in the trial area. In order to test this, a study was conducted with the following objectives: to investigate (1) Protection given by BCG and M.avium intracellulare (MAI) which is the most prevalent species, against the challenge with high and low virulent strains of M.tuberculosis in sensitised guineapigs. (2) Whether M.avium Intracellulare (MAI) interferes with the protective effect of BCG against challenge with both high and low virulent strains of M.tuberculosis. Sensitization was done with MAI in guineapigs using both oral and intradermal routes. Groups of species were immunized with BCG/Placebo and later challenged with high/low virulent strains of M.tuberculosis. Colony counts of M.tuberculosis bacilli from spleens of the animals were done to measure the protective effect.
The findings were: (1) BCG showed protection against both high and low virulent challenges. (2) MAI in both oral and intradermal routes had no effect against low virulent challenge. (3) There was no significant interaction between BCG and MAI against low virulent challenge. (4) MAI when given orally, showed a significant protection against high virulent challenge. The same was not seen with intradermal route. (5) MAI orally, interfered with the protective effect of BCG against high virulent strains of M.tuberculosis.

South Indian Variant strain of M.tuberculosis has been found to be less virulent to guineapigs through various studies from Madras and Bangalore. It was not known whether the low virulent nature of the tubercle bacilli was a fixed character or a change due to serial passages inside the body over a period of time. Hence, a study was conducted to see the behaviour of low virulent tubercle bacilli over a period of eight years during twenty passages in animals.

Ten low virulent cultures of M.tuberculosis isolated from patients belonging to Bangalore area were injected intramuscularly into guineapigs. The extent of lesions was assessed through Mitchison's Virulence scoring method. Bacilli recovered from the spleen of these guineapigs were passed into another set of animals and virulence scored. Twenty serial passages were thus performed over a period of eight years. The findings revealed that throughout the study in all the passages, the south Indian Variant of M.tuberculosis maintained its low virulent character.

Culture examination of sputum is known to be the best tool for assessing the prognosis of pulmonary tuberculosis patients. Controlled clinical trials with Short Course Chemotherapy (SCC) have shown that culture examination of sputum at the end of two months elicited a high degree of conversion, while smear results were inferior due to presence of dead organisms in the sputum. But, it is not possible to provide sputum culture facilities in the District TB Programme (DTP). Since smear examination facilities is widely available under the DTP, this paper examines the comparability of smear results with culture in patients treated with SCC. It also examines the value if any, of an early evaluation of the treatment outcome with SCC under operational conditions by doing smear examination at two months. Data from two operational studies on SCC are utilised for this purpose.

Of 256 total patients examined, 62 were smear positive at the end of two months, but 41 of them were excreting non- viable bacilli and were culture negative. Nevertheless, in predicting the final outcome of chemotherapy, no significant difference was observed between smear and culture examination at the end of two months. Smear positivity at the end of 2 months, by itself, cannot be considered to bode an unfavourable response, whereas negativity had a good probability of a favourable response. Microscopy in SCC gave as reliable a result as culture both during and at the end of chemotherapy. Hence, a properly conducted smear examination is as good as culture for diagnostic and prognostic purposes under operational conditions.

District Tuberculosis Programme (DTP) was formulated in 1962 with one of the objectives of diagnosing maximum number of tuberculosis patients. The rural health institutions implemented as Peripheral Health Institutions (PHIs) are expected to play a major role. The PHIs according to the type of facilities available, are classified into X-ray Centres (XCs), Microscopy Centres (MCs) and Referring Centres (RCs). While all centres provide treatment, XCs offer chest X-ray and sputum microscopy examination; MCs offer only sputum microscopy and from RCs sputum slides are prepared and referred for further examination to DTC/XCs/MCs. The performance of DTP activities at PHIs are collectively reported. Hence, it was worth studying categorywise performance of PHIs, which may help in developing strategy for improvement in the performance of the National Tuberculosis Programme.

OBSERVATIONS : DTP is operational in 390 (89%), out of the 438 districts in the country. Of the 17,850 implemented PHIs, 2390 (13.7%), 8717 (48.8%) and 6740 (37.8%) are functioning as XCs, MCs and RCs respectively. In all, 208 DTP reports for October to December 1993 quarter received at National Tuberculosis Institute, were analysed. Reporting efficiency of XCs, & MCs was 85%, whereas of RCs, 54%. Of the 33.1 million self reporting outpatients belonging to various PHIs, 35% attended XCs, 43% MCs and 22% RCs. Of the total sputum examinations performed during the study period, XCs examined 39%, MCs 52% and RCs 9%. Selection of chest symptomatics worked out to 1.8% for XCs, 2.0% for MCs and 0.7% for RCs. Out of the total 28,654 smear positive cases diagnosed, 56% were detected by XCs, 37% by MCs and only 7% by RCs. It is seen that XCs diagnosed 56% of the total cases by doing 39% of the total sputum examinations. The sputum positivity rate at XCs is 7.8% which is almost double that of 3.8% at the MCs and 4.4% at RCs.

Sputum Examination Efficiency (SEE) and Case Detection Efficiency (CDE) (percentage of achievement compared with expectation) have also been compared according to the category of PHIs. SEE of XCs & MCs were 70% & 78% respectively as compared to only 26.5% in RCs. The CDE of XCs, MCs & RCs were 69.1%, 36.9% and 14.1% respectively. It is observed that XCs are working satisfactorily as 35% of the total out patients attend the XCs, their reporting efficiency being 85%, sputum examination efficiency 70% and sputum positivity rate 7.8%, indicating good performance, while MCs had a low sputum positivity rate of 3.8%, suggesting that there is a large scope for qualitative improvement in Case-finding activity. While RCs cater to about 20% of the total out patients had poor performance on all account and need a great deal of technical supervision.