Prevalence surveys are useful for estimating the tuberculosis problem in different countries. Three techniques are commonly used in surveys, tuberculin test, mass miniature radiography and sputum examination. Each has its own limitations. A limitation of sputum examination is that all the sputum positive cases in the community cannot be diagnosed when only one sample of sputum is examined from each eligible person. Multiple sputum examinations are not often possible under field conditions of surveys covering the whole community. It would be worthwhile to have some idea of the extent of under-diagnosis in sputum examination. For this purpose, during an epidemiological survey, four specimens of sputum were collected within seven days of X-ray examination from each person with an abnormal chest X-ray in 30 villages of a district of south India. Each specimen was examined by Fluorescent Microscopy (FM), Ziehl Neelson (ZN) technique and culture.

There were 34 culture positive cases among 2,164 persons for whom all the four culture examination results were available. Of them, 21 (62%) were found positive on one specimen. The second specimen increased the positivity to 32 (95%). Thus, for detecting both smear and culture positive cases two specimens are adequate. A third specimen is helpful for detecting cases positive by culture alone. An estimate of prevalence obtained from one sputum specimen can be estimated for the prevalence obtained from many specimens by applying correction factor of 1.67 and estimates based on two specimens by applying 1.26. Of the remaining 37 smear positive cases detected by one specimen, 20 were smear positive and culture negative. Of the remaining 17 smear positive and culture positive, 14(82%) were detected by one smear examination only.

ZN positives not confirmed by culture (mostly with less than four bacilli reported in the smear) increased from 7 from the first specimen to 18 from all four specimens, while positives confirmed by culture method showed only a marginal increase from 13 to 15. FM did not have this disadvantage as only two were culture negative among the 18 smear positive results by FM method. Examination of two specimens by FM detected about 95% of cases demonstrable by this method. But with the ZN technique additional specimens may add more “false positives”. Thus, for detecting cases both smear and culture-positive two specimens appear adequate. A third specimen is helpful for detecting cases positive on culture only.

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.