Comparative testing of skin reactions to PPD mycobacterins from Mycobacterium tuberculosis and Mycobacterium scrofulaceum in school-age children

Comparative skin tests with 2 TU PPD-RT 23 with Tween 80 (prepared from M. tuberculosis) and 5 TU PPD-RS 95 with Tween 80 (prepared from M. scrofulaceum) were intradermally given to a total of 1,140 7-year-old children in two towns of Karviná district (340 and 255 children) and in Teplice (267 children) and Prague (278 children). In the two groups of Karviná district children the percentages of small-sized reactions (6-9 mm) to PPD-RT 23 were 13.6 and 22.3% compared to 7.1% in Teplice and 5.7% in Prague. The prevalence of small reactions to the PPD-RS 95 test in district of Karviná was 14.4 and 16.9%, in Teplice 4.5% and in Prague 6.8%. In the two towns of Karviná district the percentages of children whose reaction to PPD-RS 95 was larger than to PPD-RT 23 were 56.4 and 42.8%, in Teplice 24% and in Prague 24%. The hypothesis is advanced that the higher degree of skin hypersensitivity to the M. scrofulaceum mycobacterin which was found among the Karviná district children tested is due to sensitization with environmental mycobacteria which are common in this area.     

Prevalence of mycobacterial sensitivity in Montreal children

The prevalence of sensitivity to atypical mycobacteria was found to be low in 2152 Montreal first-grade children tested with tuberculin and one of four atypical antigens randomly allocated. Forty-nine had positive reactions to an atypical antigen. PPD-G produced more reactions than PPD-A, -B or -K. In Greater Montreal there was a higher prevalence of sensitivity to atypical mycobacteria than in the two suburban areas studied. The range of this prevalence, standardized for the type of antigen, was from 0.86 to 2.92% according to region. The highest prevalence of sensitivity to any single atypical antigen, 5.3%, was found for PPD-G in Greater Montreal.Tuberculous infection was found in 1.35% of the children. Small tuberculin reactions (5 to 9 mm to stabilized PPD 5 TU) require clarification by differential tuberculin testing. More of them are caused by M. tuberculosis than by atypical mycobacteria in this particular age group and region.Routine BCG immunization is not indicated for the particular population studied; when given to individuals at risk, its effectiveness should not be impaired by atypical sensitivity at the low level found locally. Future BCG plans require more epidemiologic data.     

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.     

A novel Candida albicans skin test antigen: efficacy and safety in man

Yeast phase Candida albicans (ATCC No. 10231) was grown in a nonantigenic medium, harvested and lyophilized. Ammonium sulfate fractions of an aqueous extract of the lyophilized cells were evaluated and the fraction yielding the highest specific delayed cutaneous reactivity in sensitized guinea-pigs was used to prepare a C. albicans skin test antigen (CASTA). The safety of the antigen was evaluated by measuring immediate and delayed (0.25, 6, 24, 48 and 72 h) cutaneous reactions in atopic and nonatopic human subjects. The outcome of three repetitive monthly Mantoux skin tests with 0.01-1 microgram antigen doses was used to test for booster effects in 14 subjects and to estimate a safe initial test antigen dose. The utility of a single skin test as a measure of cell-mediated immunity was evaluated in 40 healthy subjects. Reactor rates (greater than or equal to 2 mm, 48 h) of 40% and 85% were detected, respectively, with doses of 0.0316 and 1 microgram. Using a skin test reaction diameter greater than or equal to 5 mm at 48 h, the reactor rate was 50% for the 1-microgram dose. The only adverse reaction (45 mm, 0.25 h) was detected with the 1-microgram dose in an atopic subject who also exhibited exquisite scratch test reaginic hypersensitivity to C. albicans allergen. The prevalence of other adverse reactions to this antigen compared favorably with that to other antigens used for recall antigen testing. These studies suggest the 1-microgram CASTA dose can be used for effective, safe recall antigen skin tests.     

Eimeria stiedai: delayed hypersensitivity response in rabbit coccidiosis.

The development of delayed hypersensitivity (DH) in rabbits infected with Eimeria stiedai was shown by skin testing. A particulate antigen fraction was prepared by extraction of nonsporulated E. stiedai oocysts and found to be effective in producing dermal induration similar to that seen in a tuberculin reaction. The average diameter was 9 mm (range 7–11.0 mm, n = 26) with an average thickness of 0.4–0.5 mm for infected rabbits. All skin reactions were negative in noninfected animals (0–3.0 mm diameter and 0–0.2 mm thickness). Histological examination of dermal reactions revealed mononuclear cell infiltration within 48 hr with areas of necrobiosis. Skin reactivity was passively transferred to noninfected rabbits with lymphocyte suspensions and cell-free transfer factor but not with serum from infected skin-reactive animals. Delayed hypersensitivity was detected in 11 of 28 infected rabbits at 10 days, and by 20–30 days, 53 of the 55 animals tested showed positive skin reactions.     

Secondary and Apparent Primary Antibody Responses After Group A Streptococcal Vaccination of 21 Children

A partially purified M protein, extracted from a mouse-virulent strain of type 3, group A streptococcus, was administered subcutaneously in gradually increasing amounts at weekly intervals to 21 children in a Family Program. Seven children with type 3 bactericidal antibody in prevaccination sera showed a secondary response. Of 14 children with no detectable type 3 bactericidal antibody prior to vaccination, 13 developed definite type 3 antibody during or soon after vaccination. This response appeared to be of the primary variety in at least some of the 13 children because (i) the total incidence of antibody response (20 of 21) was greater than can be accounted for by the documented incidence of clinical and subclinical type 3 infections among children of our Family Program during a period of 14 years, (ii) the response in the 13 children with no detectable antibody prior to vaccination was more delayed than in those showing a definite secondary response, and (iii) on the average, the amount of vaccine needed for a response in these 13 children was 15 to 28 times greater than that required for the secondary response. Local skin reactions were variable among the vaccinated children. Systemic reactions were infrequent and not severe. The giving of multiple injections of partially purified M protein did not seem to cause cutaneous hypersensitivity.     

Studies on the Tuberculin Reactivity of Tuberculin-Active Peptide

A comparative study of the tuberculin potency and reactivity to TAP and PPD-S was made in humans and guinea pigs. A comparison of reactions elicited by TAP and PPD-S was made in two groups of guinea pigs previously inoculated with killed bacilli or with living bacilli. The tuberculin reactions produced by TAP and PPD-S in human beings was studied in school children from rural districts. These children were divided randomly into three groups and each group was given a different amount of TAP and PPD-S. The results obtained are as follows: (1) While relative potency of TAP was relatively much weaker than PPD-S in guinea pigs sensitized with tubercle bacilli, in human beings the tuberculin potency of TAP was approximately 1/2 or 1/3 that of PPD-S. (2) There was a marked difference in the tuberculin reactivity to TAP and PPD-S between guinea pigs immunized with killed bacilli and guinea pigs immunized with living bacilli.     

Characterization of serologic and cell-mediated reactivity of a 38-kDa antigen isolated from Mycobacterium tuberculosis

An antigen of Mycobacterium tuberculosis with an m.w. of 38,000 has been isolated by affinity chromatography using a monoclonal antibody. This antibody bound only to an antigen found in M. tuberculosis and Mycobacterium bovis BCG. The specificity of the antigen was tested in a vertical study by immunodetection on western blots reacted with hyperimmune sera against M. tuberculosis, M. bovis, and 10 other Mycobacterium species. The antigen was detected only by antisera to M. tuberculosis and M. bovis. Specificity in cell-mediated immunity was tested by skin tests in guinea pigs sensitized with M. tuberculosis, Mycobacterium intracellulare, and Mycobacterium kansasii and by lymphocyte proliferation tests. The 38-kDa antigen induced positive skin test reactions regardless of the Mycobacterium species used to sensitize the animal. The ability of the 38-kDa antigen to sensitize for cell-mediated immunity was tested by injecting mice with the 38-kDa antigen and challenging their lymphocytes in vitro with various mycobacterial antigens. Lymphocyte proliferation was observed in the presence of 38-kDa antigen, M. tuberculosis sonicate antigen, and tuberculin purified protein derivative and to M. kansasii and M. intracellulare. The 38-kDa antigen may contain a specific epitope detected by serology, but also contains epitopes that are cross-reactive for cellular immunity.     

Interpretation of the PPD skin test in BCG-vaccinated children.

Skin testing with 5 tuberculin units (TU) of purified protein derivative (PPD) of tuberculin stabilized with polysorbate (Tween) 80 was done 3 months and 1 year after immunization with bacille Calmette-Guérin (BCG) vaccine in two groups of children: one group vaccinated at birth and another group at age 6 years. Interpretation of the PPD skin test with 5 TU is possible in children 1 year and older vaccinated with BCG at birth: if the diameter of induration is more than 10 to 12 mm the reaction cannot be ascribed to BCG vaccination and is highly suggestive of supervening infection with Mycobacterium tuberculosis or occasionally atypical mycobacteria. In contrast, the interpretation of a PPD test in children vaccinated at age 6 years is extremely difficult.     

Delayed cutaneous hypersensitivity response in tetanus toxoid sensitized rhesus monkeys: predictor of anergy and value in tuberculin skin testing

The primary problem in using the tuberculin skin test in nonhuman primates is the clinical uncertainty concerning the animal's ability to elicit a delayed cutaneous hypersensitivity response. A negative tuberculin skin test can only be meaningful if the animal can produce a delayed cutaneous hypersensitivity reaction. Veterinarians deliberately sensitize animals to antigens in the form of prophylactic vaccination. Therefore, if nonhuman primates were deliberately sensitized to an antigen capable of producing a hypersensitivity response, that antigen should serve as a positive control for delayed cutaneous hypersensitivity reactions. Tetanus toxoid was chosen because repeated immunizations with this antigen is recommended routine medical practice for nonhuman primates housed outdoors. Twenty juvenile, male rhesus (Macaca mulatta) monkeys were selected for this study. The monkeys were assigned randomly to one of two groups of ten animals. The test group was vaccinated with tetanus toxoid intramuscularly at 1 month intervals for a total of three vaccinations. The control group was treated the same except saline was administered rather than tetanus toxoid. Following sensitization, the two groups of animals were challenged with tetanus toxoid intradermally. Eight of the ten monkeys in the test group responded to the tetanus toxoid while none of the control groups responded to the tetanus toxoid. Elicitation of a delayed cutaneous response in animals sensitized to tetanus antigen before challenge may serve as a positive control for delayed cutaneous hypersensitivity. This simple test may serve as a useful adjunct in making objective clinical decisions concerning anergy-suspect animals.     

Clinical Significance of Skin Reactions to Mite Extracts in Children with Asthma

The mite Dermatophagoides pteronyssinus has been identified in dust from the houses of children in Birmingham suffering from asthma. Skin tests were carried out on 150 asthmatic children with extracts of D. pteronyssinus, of the related species D. farinae, of other mites found in house dust, and of crude house dust. Though positive reactions to D. pteronyssinus were obtained more frequently and were of greater size than those to the other extracts, it was considered that D. farinae is a suitable substitute for D. pteronyssinus for skin testing.In further tests on 302 asthmatic children with mite extracts and with extracts of allergens obtained commercially reactions to the former extracts were much more common than reactions to the latter.Major skin reactions (weals with diameter of 5 mm. or more) were present in 77% of children with a history of perennial asthma and house-dust sensitivity. Hence allergy to house-dust mites, particularly D. pteronyssinus, is of considerable importance in childhood asthma, and further study of the ecology and control of the mites in dust is desirable.     

Clonal expansion of CD8+ effector T cells in childhood tuberculosis

The role of CD8(+) T cells in human tuberculosis (TB) remains elusive. We analyzed the T cell repertoire and phenotype in 1) children with active TB (< or =4 years), 2) healthy latently Mycobacterium tuberculosis-infected children, and 3) noninfected age-matched (tuberculin skin test-negative) controls. Ex vivo phenotyping of T cell subpopulations by flow cytometry revealed a significant increase in the proportion of CD8(+)CD45RO(-)CD62L(-)CD28(-)CD27(-) effector T cells (T(EF)) in the peripheral blood of children with active TB (22.1 vs 9.5% in latently M. tuberculosis-infected children, vs 8.5% in tuberculin skin test-negative controls). Analyses of TCR variable beta-chains revealed markedly skewed repertoires in CD8(+) T(EF) and effector memory T cells. Expansions were restricted to single TCR variable beta-chains in individual donors indicating clonal growth. CDR3 spectratyping and DNA sequencing verified clonal expansion as the cause for CD8(+) effector T cell enrichment in individual TB patients. The most prominent enrichment of highly similar T(EF) clones (>70% of CD8(+) T(EF)) was found in two children with active severe TB. Therefore, clonal expansion of CD8(+) T(EF) occurs in childhood TB with potential impact on course and severity of disease.     

Variation of antigenicity and serological reaction to Pneumocystis carinii in Korea.

The present study observed the variation of antigenicity of Pneumocystis carinii and serum IgG antibody reaction to the antigens from different localities in Korea. Antigens of rat P. carinii and sera of inhabitants were collected at Chunchon. Chungju, Kwangju, and Seoul during 1995-1996. Enzyme-linked Immunosorbent Assay and immunoblot were used for immune reaction. Absorbance of 1,294 human sera ranged between 0.01 and 0.93. Sera from Chunchon showed higher absorbances than those from other areas. Immunoblotting revealed IgG antibody reactions to 116, 100, and 45-55 kDa antigenic bands of rat P. carinii, but the frequencies of positive reaction to individual bands were variable by localities. Total 62.6% of the sera showed the reaction to 116 kDa band while 37.7% reacted to 100 kDa band and 32.0% to 45-55 kDa bands. For the reaction to 116 kDa, the reaction rate was 60.0% to 82.6% by localities. It is found that the reaction rates of the human sera to rat P. carinii antigen are variable according to the localities. Also, the high molecular antigen of 116 kDa of rat P. carinii is the most frequent antigenic band reacting to human sera.     

利用rCFP-10/ESAT-6融合蛋白刺激γ-IFN体外释放测定替代结素皮试检出结核感染的可行性

评估rCFP-10/ESAT-6融合蛋白刺激γ-IFN体外释放测定与结素皮试检出结核感染的敏感性及特异性。对疑似结核病患者共229例进行随机、双盲、平行、对照、前瞻性试验,后经细菌培养证实患结核病的病人共129人,没有结核病史的非结核病患者共100人。以某一特定的γ-IFN体外释放水平及结素皮试反应硬结直径?10 mm为阳性切割值,rCFP-10/ESAT-6融合蛋白刺激γ-IFN体外释放测定的敏感性为96%,显著高于结素皮试(89%)(χ2=4.92;0.025相似文献   

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.     

Epidemiological aspects of toxoplasmosis in schoolchildren residing in localities with urban or rural characteristics within the city of Rio de Janeiro, Brazil

Immunofluorescence tests (IF) for toxoplasmosis were performed on a total of 608 schoolchildren in elementary and junior high grades. 166 being in the Bonsucesso district (an urban region of Rio de Janeiro) and 442 children from locations within the lowlands of Jacarepaguá (with rural characteristics). All the IF-IgM were nonreactive, whilst 416 schoolchildren (68.4%) were IF-IgG serum-reactive (greater than or equal to 1:16). The percentages of serum-reactives in Jacarepaguá were significantly higher than in Bonsucesso, both as regards the total number of schoolchildren (p less than 0.001), as also when subdivided according to the age-grades from six to eight years (p less than 0.001) or from twelve to fourteen (p less than 0.05). Both in Jacarepaguá and in Bonsucesso, the prevalence of reactions in the 12 to 14 year age-grade was significantly greater than in the 6 to 8 year age-grade (p less than 0.001 in both cases). Expressively larger prevalences of serum reactions were found in Jacarepaguá among schoolchildren who preferred eating raw or undercooked meat, as well as among those having cats as pets; this occurred equally in the 6 to 8 year and in the 12 to 14 year age-grades. In Bonsucesso, the only significant difference was in the 6 to 8 year age-grades that had cats as pets. Thus, it has been verified that the risk of infection is greater and more precocious in localities with rural characteristics than in urban regions.     

Antigen recognition by serum antibodies in non-human primates experimentally infected with Mycobacterium tuberculosis

Tuberculosis is a significant threat to non-human primates and their caretakers. The diagnosis of tuberculosis in living non-human primates is currently based on the tuberculin skin test, which is cumbersome and sometimes inaccurate. Development of an accurate serodiagnostic test requires identification of the key antigens of Mycobacterium tuberculosis involved in antibody production. When sequential serum samples obtained from 17 cynomolgus, rhesus, and African green monkeys up to seven months since experimental infection with M. tuberculosis Erdman were screened for antibody against purified proteins of M. tuberculosis, three highly seroreactive antigens were identified. One protein, ESAT-6, reacted with sera from all infected animals. Two additional proteins, alpha-crystallin and MTSA-10, were recognized by sera from approximately 90% of infected animals. Time course analysis of antibody production indicated that the earliest response was usually to ESAT-6 alone or to ESAT-6 and other antigen(s). These results provide experimental evidence of the potential value of ESAT-6 as an antigen for use in serodiagnosis of tuberculosis in non-human primates.     

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.     

Cell-mediated response to BCG investigated by leukocyte migration test from an agarose droplet

The modified leukocyte migration test (LMT) from an agarose droplet with the antigen stimulation (by BCG) is proposed in the present work. The BCG concentrations ranging from 1.25 up to 130 mg/l were used to examine 20 tuberculin (PPD) skin test negative and 10 PPD positive patients, which suffered from lung diseases. The optimal concentrations were 6.25 and 25.0 mg/l. The mean index values of a 20 membered control group ranged from 0.7 up to 0.88 when stimulated with the lower BCG concentration, and they were of 0.53 up to 0.69 at higher BCG concentration. In spite of its suitable indicatory properties as to ascertain cell mediated immunity state, the LMT with the BCG were of no use as a tuberculin skin test correlate.     

Colonization with nontuberculous mycobacteria is associated with positive tuberculin skin test reactions in the common marmoset (Callithrix jacchus)

Mycobacterium tuberculosis infections can result in significant morbidity and mortality in nonhuman primate colonies. Preventative health programs designed to detect infection routinely include tuberculin skin testing (TST). Because Mammalian Old Tuberculin used for TST contains antigens common to a variety of mycobacterial species, false-positive results can occur in animals sensitized to nontuberculous mycobacteria (NTM). Over 11 mo, a large colony of common marmosets (Callithrix jacchus) demonstrated a 3.6% prevalence of equivocal or positive TST reactions (termed 'suspect reactions'). Culture of gastric aspirates, bronchoalveolar lavage fluid, and feces revealed a single animal with a positive fecal culture for Mycobacterium gordonae. PCR amplification of M. gordonae DNA in feces collected from animals with suspect TST reactions (demonstrating a 66.7% colonization rate) and colony controls (demonstrating a 14.3% colonization rate) revealed a significant association between suspect TST reactions and intestinal colonization. Gross and histopathologic evaluation revealed a multifocal lymphadenopathy and granulomatous lymphadenitis in 2 of 4 TST-positive marmosets examined. Counter to expectations, granulomatous lymphoid tissue was culture-positive for M. kansasii rather than M. gordonae. Detection of M. gordonae in the feces of TST-suspect animals likely represents an apathogenic intestinal colonization that may serve as an indicator of NTM exposure, whereas evidence of histopathologic disease is associated with the more pathogenic M. kansasii. Although a high index of suspicion for M. tuberculosis should always be maintained, colonization with NTM organisms represents a cause of suspect TST reactions in common marmosets.