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Although the tuberculin test has aided in the diagnosis of tuberculosis for more than 85 years, its interpretation is difficult particularly because sensitization with non-tuberculous mycobacteria leads to false positive tests. Using the guinea pig model of tuberculosis, we have recently described a recombinant antigen (DPPD) that could circumvent this problem. The DPPD gene is unique to the M. tuberculosiscomplex organisms and is absent in the organisms representative of all other members of the Mycobacteriumgenus. Moreover, DPPD induced strong DTH in 100% of the guinea pigs infected with M. tuberculosisand in none of the guinea pigs immunized with nine different species of Mycobacterium. Here we present results of a clinical investigation using DPPD. Mantoux test using both PPD and DPPD was initially performed in 26 patients with confirmed pulmonary tuberculosis and in 25 healthy PPD negative individuals. The results indicated that both PPD and DPPD elicited DTH in 24 out of the 26 patients. No DTH was observed in any of the PPD negative individuals. In addition, a small clinical trial was performed in a population of 270 clinically healthy and randomly selected individuals. DPPD produced a bimodal histogram of skin reaction size and PPD produced a skewed histogram. Because the DPPD gene is not present in non-tuberculous bacilli, these results suggest that this molecule can be an additional tool for a more specific diagnosis of tuberculosis.

The efficacy of GM-CSF was investigated in 20 neutropenic patients (< 1500 neutrophils/µl) with acute visceral leishmaniasis due toLeishmania chagasi. Patients were randomized to receive either GM-CSF, 5 µg/kg daily (intravenously or subcutaneously), or placebo for ten days, in combination with pentavalent antimony, 10–20 mg/kg daily for 20 days. Neutrophil counts were significantly greater on days 5 and 10 of treatment in the GM-CSF group compared with the placebo group (p<0.02). Eosinophil and monocyte counts were also significantly increased in the GM-CSF group at day 10 (p≤0.03). Interestingly, at day 30, platelet counts were significantly higher in the GM-CSF treated group (p=0.007). Haemoglobin levels were significantly increased in the GM-CSF group on days 5 and 10 (p=0.04 and 0.02, respectively). Patients in the GM-CSF group experienced fewer secondary bacterial or viral infections than placebo patients. Infections occurred in only three patients given GM-CSF compared with eight patients given placebo (p<0.04). All patients had complete resolution of disease symptoms at three months. Few adverse events were recorded. GM-CSF given subcutaneously at a dose of 5 µg/kg daily for ten days was well tolerated, reversed neutropenia rapidly and reduced the number of secondary infections in patients with leishmaniasis.

The diagnosis of visceral leishmaniasis (VL), a serious and often fatal parasitic disease caused by members of the Leishmania donovani complex, remains problematic. Current methods rely on clinical criteria, parasite identification in aspirate material, and serology. The latter methods use crude antigen preparations lacking in specificity. A previously described cloned antigen, rK39, of Leishmania specific for all members of the L. donovani complex (L. chagasi, L. donovani, L. infantum) was very useful in the serodiagnosis by ELISA of both human and canine VL. The present study demonstrated that rK39 seroreactivity correlated with active disease. The sera from early or self-healing infected subjects reacted with leishmanial lysate and were generally nonreactive with rK39. These data demonstrate the utility of rK39 in the serodiagnosis of VL and as an indicator of active disease.

High tumor necrosis factor-α (TNFα) levels were present in the serum of24 of28 active visceral leishmaniasis (VL) patients (142.9 ± 113.9pg/ml, mean ± SD), whereas levels were not elevated in 26 of 30 patients with cryptic leishmanial infection (16 asymptomatic, 4 with self-healing subclinical infection, and 10 posttreatment VL cases). Serum TNFα levels were also not elevated in 15 normal volunteers (11.3 ± 15.6 pg/ml) and in 10 patients with tegumentary leishmaniasis (19.1 ± 10.8pg/ml). Leishmanial infection of human monocyte-derived macrophages enhanced the basal TNFα production by these cells, and this effect was further potentiated by treatment with recombinant interferon-γ, After effectivetreatment ofVL patients, serumTNFα levelsdropped rapidly (129 ± 112 vs. 9 ± 13 pg/ml in 10 days), even before clinical parameters such as spleen size or parasitism, white blood cell count, or levels of hemoglobin returned to normal values. On the other hand, patients unresponsive to treatment remained with elevated levels (276 ± 69 vs. 155 ± 71 pg/ml in 10 days). Thus, serum TNFα levels in VL patients are a good parameter to monitor in determining host response to therapy.

The epidemiology, clinical patterns, and risk factors for visceral leishmaniasis were prospectively studied in an endemic area of Brazil. The prevalence of disease was 3.1% for children <15 years of age, and the annual incidence was 4.3 cases per 1,000 children. The number of children with disease fluctuated yearly and seasonally, and distribution of the disease varied within the endemic area. Risk factors included young age (median, three years) and malnutrition before the onset of disease. Intestinal parasitism, recent migration into the area, and house location within the area did not influence the progression of infection to disease. Serological testing indicated that 7.5% of children were infected with Leishmania each year and that the ratio of disease to infection was 1:18.5 for the whole area and 1:6.5 for the section with the highest prevalence of disease. Early diagnosis and therapy altered clinical patterns of the disease.