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Studies were undertaken in three independent centers to compare the newer, in vitro radiometric macrophage (M phi) assay with the conventional mouse foot pad infection for the diagnosis of dapsone resistance. Results obtained on 12 bacilliferous patients showed good concordance in both assays. One strain diagnosed as sensitive in the M phi assay was found to be resistant in the mouse foot pad. Three Mycobacterium leprae strains considered to be partially resistant in the mouse infection behaved as resistant strains in the M phi cultures. Attention is drawn to a rapid in vitro method for the identification of drug-resistant bacilli in leprosy patients.

Mycobacterium leprae cells extracted from the skin biopsies of 14 bacilliferous lepromatous patients were maintained in human-murine macrophage cultures for 3 weeks in the presence of [3H]thymidine and DDS (4,4'-diaminodiphenyl sulfone). All cultures except one containing freshly extracted viable bacilli showed significant incorporation of [3H]thymidine as compared to control cultures containing heat-killed bacilli of the corresponding strain. Six susceptible strains of M. leprae obtained from untreated, freshly diagnosed patients showed significant inhibition of the uptake of the radiolabel in the presence of 3 and 10 ng of DDS per ml per culture. Eight strains of M. leprae obtained from patients clinically suspected of DDS resistance were tested in a similar manner. These strains were also concurrently inoculated in the footpads of mice given orally 10(-2), 10(-3), and 10(-4) g of DDS per 100 g of body weight for 9 months. Concordant results were obtained by both methods: five strains were found to be resistant, one was susceptible, and one was partially resistant. Strain VIII did not incorporate [3H]thymidine in the macrophage cultures and proved to be resistant in the mouse footpad. The macrophage culture system provides a sensitive, rapid screening method for the early diagnosis of DDS resistance.

A case of borderline lepromatous leprosy with a history of 5 years duration of disease, was first seen in 1971 and treated with graded doses of dapsone in the OP Clinic of the Institute. He became inactive and bacteriologically negative in 3 years. While continuing on dapsone therapy he relapsed into active dapsone resistant leprosy 3 1/2 years later. He was admitted in the hospital and given Rifampicin 600 mg. daily for 15 days along with dapsone 100 mg. daily. He again became inactive and bacteriologically negative within 3 years. 3 years later under regular dapsone therapy he relapsed again for the second time into active BT leprosy, but remained bacteriologically negative. He was given 3 drug regimen subsequently and became clinically inactive within 15 months.

This paper describes a microculture rapid assay using radiolabeling and mouse macrophages to determine the viability and the drug susceptibility or resistance of Mycobacterium leprae. Comparison of M. leprae resident macrophage cultures maintained in 96-well flat-bottomed plates showed results for viability and susceptibility or resistance to dapsone that were similar to results for concurrent cultures in Leighton tubes with greater numbers of bacilli and macrophages.

The Central Leprosy Teaching and Research Institute (C.L.T.R.I.) Chengalpattu, took up studies on dapsone resistance in M. leprae from 1974. From 1978, the study was further strengthened by a project under THELEP (TDR) for eliciting information on the efficacy of certain drug regimens. The Thelep studies were to be conducted only on the dapsone sensitive untreated cases and, therefore, directed towards the detection of primary resistance, while the non-THELEP institutional studies were concentrated on secondary dapsone resistance problem. These two studies together detected 99 cases of dapsone resistance in the patients who attended CLTRI Hospital during 1974-81; 23 of them, were of primary origin, 16, 6 and 1 showing mild (RI), moderate (RII), and high (RIII) grades respectively. Of the remaining 76 cases of secondary resistance, 7 and 69 were of RII and RIII grades respectively. The need for vertical and horizontal monitoring of the drug resistance problem has been pointed out.