Vaccination with Freeze-Dried BCG

Sixty first-year medical students, negative to 1:2000 and 1:100 dilutions of old tuberculin (OT), were vaccinated with freezedried BCG vaccine. At six months, 98.3% were found to be positive reactors as ascertained by the intracutaneous (Mantoux) method, using 1:2000 and 1:100 dilutions of OT, and 73.7% as ascertained by the Heaf test, using a tuberculin PPD solution containing 100,000 T.U. per ml.     

Tuberculin Test in Children with Malnutrition

Tuberculin tests with Heaf''s multiple-puncture method, as well as intradermal tests with varying strengths of old tuberculin, were carried out on 402 children with severe malnutrition. From the radiological and bacteriological findings 51 children (12.5%) were considered to have active tuberculosis. The Heaf test was positive in only 11 of these children, but the intradermal test using 100 tuberculin units was positive in a further 18. This confirms previous findings that tuberculin sensitivity is impaired in malnourished children, and suggests that a higher dose of tuberculin is more likely to elicit a positive response.     

Comparison of the tine and Mantoux tuberculin tests. Report of the Tuberculin Subcommittee of the Researched Committee of the British Thoracic Association.

Intracutaneous (Mantoux 5 TU) and tine tuberculin tests were performed on the opposite arms of 307 people. The results of each test were read by two independent observers at either 48 or 72 hours. Positive readings were recorded in 59% of the Mantoux tests; induration was 10 mm or more in 34.7% of cases. Positive readings were recorded for 3.9% of the tine tests, and a further 15.5% were recorded in the doubtful category. The tine test is unsuitable for epidemiological use because of the high proportion of negative and doubtful results in people positive on the Mantoux test. For the same reasons its usefulness in clinical practice is very limited.     

Interpretation of Positive Reactions to Heaf Tuberculin Test in London Schoolchildren

A study of 289 children aged 12-14 years found to be Heaf test positive at routine testing was made to determine the likely cause of the positive tests. Only 12% of children grade 1 Heaf positive were likely to have had a previous infection with Mycobacterium tuberculosis, 49% were likely to have had infections with atypical mycobacteria, and in 39% no cause for the positive Heaf test was found.The positive Heaf test seems to us to be too sensitive. Both a negative and grade 1 positive should be regarded as not significant and these children given B.C.G. vaccine. Heaf grades 2, 3, and 4 should be regarded as significant and the children investigated for M. tuberculosis infection.     

Comparison of Mantoux and Tine Tuberculin Skin Tests in BCG-Vaccinated Children Investigated for Tuberculosis

Background

Tuberculin skin tests (TSTs) are long-established screening methods for tuberculosis (TB). We aimed to compare agreement between the intradermal Mantoux and multipuncture percutaneous Tine methods and to quantify risk factors for a positive test result.

Methodology/Principal Findings

1512 South African children younger than 5 years of age who were investigated for tuberculosis (TB) during a Bacille Calmette Guerin (BCG) trial were included in this analysis. Children underwent both Mantoux and Tine tests. A positive test was defined as Mantoux ≥15 mm or Tine ≥ Grade 3 for the binary comparison. Agreement was evaluated using kappa (binary) and weighted kappa (hierarchical). Multivariate regression models identified independent risk factors for TST positivity. The Mantoux test was positive in 430 children (28.4%) and the Tine test in 496 children (32.8%, p<0.0001), with observed binary agreement 87.3% (kappa 0.70) and hierarchical agreement 85.0% (weighted kappa 0.66). Among 173 children culture-positive for Mycobacterium tuberculosis, Mantoux was positive in 49.1% and Tine in 54.9%, p<0.0001 (kappa 0.70). Evidence of digit preference was noted for Mantoux readings at 5 mm threshold intervals. After adjustment for confounders, a positive culture, suggestive chest radiograph, and proximity of TB contact were risk factors for a positive test using both TST methods. There were no independent associations between ethnicity, gender, age, or over-crowding, and TST result.

Conclusions/Significance

The Tine test demonstrated a higher positive test rate than the Mantoux, with substantial agreement between TST methods among young BCG-vaccinated children. TB disease and exposure factors, but not demographic variables, were independent risk factors for a positive result using either test method. These findings suggest that the Tine might be a useful screening tool for childhood TB in resource-limited countries.     

Evaluation of a Tuberculosis Mass Survey Conducted in the City of Chatham and in Kent County,Ontario, in 1963

Computer analysis of the results of a mass tuberculosis survey of the population of Chatham city and Kent County, Ontario, was carried out to assess the efficacy of such surveys as a case-finding device. Six cases of active tuberculosis were found during the survey and approximately 50% of the 89,427 inhabitants of Chatham and Kent County were surveyed with radiographs and Heaf skin tests.A follow-up study of the Heaf test conducted in the public school of Wallaceburg detected 70 Heaf-positive children out of a total of 1190. These Heaf-positive children were further tested with a Mantoux test (5 T.U.) and chest radiography. Only 11 children of the original 70 were Mantoux-positive.These findings further support the hypothesis that the Heaf test, as now read, over-reads for M. tuberculosis infections.     

Assessment of tine tuberculin test.

Mantoux (10 tuberculin units) and tine tuberculin tests were performed in 190 subjects and the results recorded at 48, 72, and 96 hours. Altogether 1010 readings were recorded. Of these, 555 (55%) were Mantoux-test positive and 525 (52%) tine-test positive. While the tine test is marginally less sensitive, its advantages are simplicity and ease and speed of application. It thus provides a useful screening test for eliciting tuberculin sensitivity.     

Tuberculin Reactions in Bulls And Boars Sensitized with Atypical Mycobacteria from Sawdust

Due to a planned export from a combined bull and boar station, more than 70 boars and 100 bulls were examined by tuberculin tests. Distinct reactions to avian tuberculin appeared in about half of the animals. Many of them also reacted to bovine tuberculin. For diagnostic purposes, many of the reactors were slaughtered, and samples from these and from the environment were examined bacteriologically. Strains of Mycobacterium avium were isolated from only 2 out of 14 reacting boars and from none of the 23 reacting bulls. No isolation of Mycobacterium bovis was made. However, atypical mycobacterial strains, classified as Runyon Group III and IV, were isolated from 3 boars, 2 bulls, 1 pigeon and from many samples of sawdust. The isolation of identical fast-growing mycobacterial strains from the sawdust used in the pens for the reacting boars and bulls, was especially remarkable. The strains differed enzymatically and biochemically from those isolated from other sources. This indicated a possible sensitization of the animals with similar mycobacterial strains. Possible cross-reactions to avian and bovine tuberculin were investigated in tuberculin assays with guniea pigs and pigs sensitized to one of the mycobacterial strains isolated. Distinct reactions to both avian and bovine tuberculin appeared in all the animals. From these results it was concluded that the tuberculin reactions in the boars and the bulls were not due to any tuberculous infection in the herd, but to a sensitization of the animals with atypical mycobacteria.     

Tuberculin Activity of Mycobacterial Fractions Obtained by Chromatography

Commercial purified protein derivatives (PPD), old tuberculin (OT), the bacillary extract, and the culture filtrate of Mycobacterium tuberculosis H37Ra were submitted to Sephadex G-25 and diethylaminoethyl (DEAE)-cellulose chromatography. The ability of the fractions obtained to elicit delayed dermal hypersensitivity in M. tuberculosis H37Ra-infected guinea pigs was studied. Skin tests with Sephadex fractions in M. tuberculosis H37Ra-infected guinea pigs showed that the tuberculin activity was localized in the first fraction. All other Sephadex fractions were nonessential and nonspecifically irritating. Fractions from chromatography of Sephadex G-25 fraction 1 on DEAE-cellulose columns showed that all but the first were able to elicit delayed hypersensitivity reactions. There was a variability in the capacity to elicit the tuberculin reaction according to the fraction injected and the stage of tuberculous infection in guinea pigs. Compared to the others, the seven lots of commercial PPD were variable in composition and content. They contained both essential and nonessential materials for the tuberculin reaction. Sephadex fraction 1 would appear to be a better tuberculin as it excludes nonessential nonspecifically irritating elements and contains the complement able to elicit the tuberculin reaction. Its methodological simplicity would be economically advantageous.     

The Tuberculin Test

A brief historical review of the tuberculin test is followed by consideration of the immunological factors concerned in the tuberculin reaction. The problems in the interpretation of the test, the dosage and types of material used, and methods of administration are discussed. The current problem of tuberculin reversion with and without treatment, as exemplified by the current study of Halifax school children, is considered in some detail. The role of the tuberculin test in the present-day program of tuberculosis control is indicated.     

Leukocyte procoagulant activity in man: an in vitro correlate of delayed-type hypersensitivity

Human mononuclear leukocytes generate cell-bound procoagulant activity (LPCA) after incubation with an antigen (mumps or tuberculin) to which the donor was previously sensitized. An inhibitor of coagulation appears to be liberated into the extracellular culture fluid during incubation of leukocytes with the sensitizing antigen. Removal of this activity before measuring LPCA resulted in a reliable test that correlated directly with delayed skin reactivity. The assay was particularly sensitive in that cells from weakly sensitized donors who reacted only to high doses of tuberculin (100 TU) in the delayed skin tests produced detectable LPCA in vitro. By contrast cells from weakly sensitized donors did not react to PPD in the lymphocyte blast transformation test or the direct macrophage migration inhibition factor test. The LPCA assay correlated closely with the blast transformation and MIF tests in which cells were used from more strongly sensitized donors who reacted in skin tests with lower doses of tuberculin (1 or 10 TU). The assays were antigen-specific in that cells from donors sensitive to mumps antigen but not to tuberculin reacted only to mumps antigen in vitro. The assay was extremely reproducible; cells from individual donors reacted to the same extent over a period of 8 mo). We propose that the assay system reported here represents an improved method for the measurement of cell-mediated immunity in vitro because it requires fewer donor cells, is technically simpler, and is more sensitive than previously described methods.     

Retention of 14C-Labeled Tuberculin Purified Protein Derivative in the Skin of Sensitized and Nonsensitized Animals

Tuberculin purified protein derivative labeled with (14)C ([(14)C]PPD) with a biological potency equivalent to the International Standard for tuberculin PPD was used to study the retention of tuberculin PPD in the skin of sensitized and nonsensitized animals. We found that [(14)C]PPD was almost entirely cleared from the skin test site during the first 18 to 24 h after injection and that when approximately 5% of the initial concentration of [(14)C]PPD was present in the skin test site, the size of the tuberculin skin reaction in sensitized guinea pigs was at its maximum. Furthermore, the addition of 5 or 50 mug of Tween 80 per ml to a solution of PPD did not change either the rate of clearance of PPD from the skin test sites of sensitized guinea pigs or the size of the tuberculin skin reactions. There was no difference in the rate of clearance of [(14)C]PPD from the skin test sites between sensitized and nonsensitized guinea pigs and between guinea pigs of different age. However, there was a significant difference in the rate of clearance of [(14)C]PPD between the guinea pig and the mouse. Finally, the percentage of [(14)C]PPD retained in the site of injection at 24 h was in the neighborhood of 5% of the initial concentration of the solution of PPD injected. The significance of these phenomena is discussed.     

Skin reactivity to mycobacterial antigens in children living in an area of Mycobacterium xenopi endemicity

Intradermal skin tests with a 2TU dose of PPD-RT 23 prepared from M. tuberculosis and 0.1 ug/0.1 ml of PPD-RS 631 from M. xenopi were simultaneously carried out in 378 7-year-old children from two localities in North-Bohemian region's capital Ustí n. Lab., a focus of M. xenopi endemicity repeatedly confirmed since its disclosure in 1980 by positive M. xenopi isolations from humans and public water supply network. A further group 157 children serving as controls was from Prague district 4 where no presence of M. xenopi strains was ever recorded. All of these children had received routine immunization at birth with Czech BCG vaccine. The children from the two endemic localities were found to give a positive 6 mm or greater reaction to M. xenopi mycobacterin in 43.3% and 22.3%, to human tuberculin in 12.8% and 12.6%, respectively. The frequency histogram clearly separated a group of reactors with 8-18 mm indurations from a group of nonreactors showing a skin induration of 4-8 mm. The higher reactivity of this exposed child population was also reflected in a larger proportion of reactions greater to M. xenopi PPD than to human tuberculin antigen: the reactions greater by 1-5 mm accounted, respectively, for 25.1% and 20.6%, reactions greater by 6 mm or more for 23.7% and 15.9%. Among a group of children from Prague district 4, 6.4% had medium-sized and 3.8% large-sized reactions to M. xenopi antigen; the proportion of reactions greater to M. xenopi antigen than to human tuberculin accounted for only 5.1%, reactions greater to tuberculin than to sensitin were here in slight predominance. The evidenced skin sensitization to M. xenopi mycobacterin is suggested to result from the different degrees of exposure to infection by environmental mycobacteria.     

Lymphocyte Stimulation and Leucocyte Migration Inhibition as Diagnostic Tools for the Detection of Mycobacterium Avium Infection in Cattle

The tuberculin skin test is the conventional method of detecting infections with mycobacteria in animals. A positive reaction is considered to reflect cell-mediated immunity (CMI). CMI against mycobacteria can be studied by in vitro systems using suspensions of blood lymphocytes or leucocytes. The reactivity of these cells to different antigens can be measured in the lymphocyte stimulation (LS) (Muscoplat et al 1975, Bergman 1976, Johnson & Morein 1976), or leucocyte migration inhibition (LMI) (Aalund 1970, Clausen 1973) tests.     

Comparative study of RT23 and IC-65 tuberculins tested on children with tuberculosis

This study, conducted in 2009, proposed to evaluate and compare the biological potency of two different tuberculins, RT23 (Statens Serum Institute, Copenhagen) and IC-65 (Cantacuzino Institute, Bucharest) when administered to 89 children with confirmed tuberculosis, admitted to Paediatric Department of Pneumophtysiology Institute, Bucharest. Mean age of subjects was 10.4 years [SD (standard deviation) = 5.2 years; variance = 27.2], and sex distribution in the group was: 55.1% girls and 44.9% boys. Tuberculin skin tests were performed using Mantoux method simultaneously with the two tuberculins in the same concentration, 2TU (tuberculin units)/0.1 ml. RT23 skin test reactions ranged from 8 mm to 18 mm (mean = 12.8 mm, SD = 2.1 mm, variance = 4.4; median = 12.0), and IC-65 reactions ranged from 8 mm to 18 mm (mean = 13.1 mm; SD = 2.1 mm; variance = 4.3; median = 13.0). The mean difference in paired reaction sizes for the two reagents was 0.04 mm and was not statistically different from zero (P value = 0.3). The difference in reaction sizes was = 2 mm in 70.8% and = 5 mm in 7.9% patients. With a cutoff of 10 mm to define a positive reaction, the results were highly correlated with a sensitivity of 98.9% for RT23 and 97.8% for IC-65. No statistically significant difference was established for the efficacy of the two commercially available PPD TST reagents, both tuberculins appearing to have equivalent potency.     

Immunologic monitoring and immunotherapy in Ewing's sarcoma

Summary Serial immunological monitoring was performed on 31 patients with Ewing's sarcoma who were on a randomized immunotherapy trial with BCG administered by dermal scarification with a Heaf gun. Patients were skin-tested for delayed hypersensitivity reactions (DCHR) to recall antigens and extracts of tumor cells, and with keyhole limpet hemocyanin (KLH). In vitro testing consisted of lymphocyte counts, percentages of cells forming rosettes with sheep erythrocytes at 29° C and at 4° C, and leukocyte migration inhibition to tuberculin (PPD) and to 3 M KCl extracts of tumor cells. At the time of diagnosis, nearly all patients had positive DCHR to mumps and streptococcal antigens and were negative to PPD. Neither the skin tests nor the lymphocyte counts at this time gave useful prognostic information. In tests during and after therapy, the patients who responded and remained free of detectable disease had a higher incidence of DCHR to KLH and of rosette values in the normal range than did the patients who developed recurrent disease. The BCG immunotherapy had no apparent effect on immunologic parameters except for conversion of reactions to PPD.     

Prevalence of Mycobacterium tuberculosis infection among injection drug users in Toronto

BACKGROUND: Injection drug users are at increased risk of Mycobacterium tuberculosis infection and active tuberculosis (TB). The primary objective of this study was to determine the prevalence of M. tuberculosis infection among injection drug users in Toronto, as indicated by a positive tuberculin skin test result. An additional objective was to identify predictors of a positive skin test result in this population. METHODS: A cross-sectional study was carried out involving self-selected injection drug users in the city of Toronto. A total of 171 participants were recruited through a downtown Toronto needle-exchange program from June 1 to Oct. 31, 1996. RESULTS: Of 167 subjects tested, 155 (92.8%) returned for interpretation of their skin test result within the designated timeframe (48 to 72 hours). Using a 5-mm cut-off, the prevalence rate of positive tuberculin skin test results was 31.0% (95% confidence interval 23.8% to 38.9%). Birth outside of Canada and increasing age were both predictive of a positive result. INTERPRETATION: There is a high burden of M. tuberculosis infection in this population of injection drug users. The compliance observed with returning for interpretation of skin test results indicates that successful TB screening is possible among injection drug users.     

Incidence of tuberculosis and the predictive value of ELISPOT and Mantoux tests in Gambian case contacts

Background

Studies of Tuberculosis (TB) case contacts are increasingly being utilised for understanding the relationship between M. tuberculosis and the human host and for assessing new interventions and diagnostic tests. We aimed to identify the incidence rate of new TB cases among TB contacts and to relate this to their initial Mantoux and ELISPOT test results.

Methods and Findings

After initial Mantoux and ELISPOT tests and exclusion of co-prevalent TB cases, we followed 2348 household contacts of sputum smear positive TB cases. We visited them at 3 months, 6 months, 12 months, 18 months and 24 months, and investigated those with symptoms consistent with TB. Those who were diagnosed separately at a government clinic had a chest x-ray. Twenty six contacts were diagnosed with definite TB over 4312 person years of follow-up (Incidence rate 603/100,000 person years; 95% Confidence Interval, 370–830). Nine index and secondary case pairs had cultured isolates available for genotyping. Of these, 6 pairs were concordant and 3 were discordant. 2.5% of non-progressors were HIV positive compared to 12% of progressors (HR 6.2; 95% CI 1.7–22.5; p = 0.010). 25 secondary cases had initial Mantoux results, 14 (56%) were positive ; 21 had initial ELISPOT results, 11 (52%) were positive; 15 (71%) of 21 tested were positive by one or the other test. Of the 6 contacts who had concordant isolates with their respective index case, 4 (67%) were Mantoux positive at recruitment, 3 (50%) were ELISPOT positive; 5 (83%) were positive by one or other of the two tests. ELISPOT positive contacts, and those with discordant results, had a similar rate of progression to those who were Mantoux positive. Those negative on either or both tests had the lowest rate of progression.

Conclusions

The incidence rate of TB disease in Gambian TB case contacts, after screening for co-prevalent cases, was 603/100,000 person years. Since initial ELISPOT test and Mantoux tests were each positive in only just over half of cases, but 71% were positive by one or other test, positivity by either might be the best indication for preventive treatment. These data do not support the replacement of the Mantoux test by an ELISPOT test in The Gambia or similar settings.     

Correlation of tuberculin-induced lymphocyte transformation with skin test reactivity and with clinical manifestations of sarcoidosis

In vitro lymphocyte reactivity to tuberculin (PPD) was studied in buffy coat cultures from 87 patients with sarcoidosis and from 64 controls. A strong correlation was found between PPD-induced lymphocyte transformation and skin reactivity. No significant differences were found in the in vitro response of lymphocytes from skin test positive patients with sarcoidosis and from controls with the same degree of skin test reactivity. In patients with sarcoidosis negative to 100 TU, tuberculin sensitivity could be demonstrated in vitro significantly more often than in comparison subjects. Both in vivo and in vitro tuberculin sensitivity and “spontaneous” transformation were significantly more frequent in patients with erythema nodosum.