- Email: [email protected]
Performance of QuantiFERON-TB Testing in a Tuberculosis Outbreak at a Primary School
Performance of QuantiFERON-TB Testing in a Tuberculosis Outbreak at a Primary School PAOLA MOLICOTTI, PHD, ALESSANDRA BUA, PHD, GRAZIELLA MELA, MD, PAOLINA OLMEO, MD, RENZO DELOGU, MD, SILVIA ORTU, PHD, LEONARDO ANTONIO SECHI, PHD, AND STEFANIA ZANETTI, PHD
This study compared the effectiveness of the QuantiFERON-TB Gold (QFT) assay with the Mantoux tuberculin skin test to detect Mycobacterium tuberculosis infection in 29 children during a school outbreak of tuberculosis. Of the 21 children with M tuberculosis infection, 11 had a radiograph suggestive of the infection. The QFT assay was positive in all 21 of the children, and the Mantoux test was negative at first testing in 2 children (1 of whom was the sentinel case). The findings demonstrate that the QFT test is extremely useful in accurately identifying infected and uninfected children, permitting rapid intervention. (J Pediatr 2008;152:585-6)
iagnosis of tuberculosis (TB) in children is based on clinical examination, chest radiography, and the Mantoux test.1 Until recently, despite its limitations,2 the Mantoux test was the only tool available for the diagnosis of latent tuberculosis infection (LTBI). QuantiFERON-TB Gold (QFT) (Cellestis Limited, Carnegie, Victoria, Australia) is an in vitro immunologic assay that measures the interferon-gamma released by T lymphocytes sensitized with the Mycobacterium tuberculosis–specific antigens ESAT-6 and CFP-10. The specificity and sensitivity of this assay has been demonstrated in adults, and excellent results have been obtained for both active TB3 and LTBI.4 In early summer 2006, a 10-year-old child was admitted to the Pediatric Infectious Diseases Clinic of the University of Sassari with fever, cough, and a chest radiograph compatible with a primary TB complex. Subsequently, the Mantoux and QFT tests were performed. M tuberculosis was isolated in culture from the gastric aspirate, and the molecular test was positive for M tuberculosis as well. Immediate contact investigation (chest radiograph, Mantoux, QFT) was performed for all of the child’s fifth-grade classmates and for other children in different classrooms attending the same school. This outbreak of TB prompted us to investigate the effectiveness of the QFT assay and to compare the results with the Mantoux test as a tool for diagnosing TB in children.
D
METHODS A total of 29 children (16 males and 13 females age 6 to 10 years, 19 of whom were the sentinel case’s classmates), were examined. None of the children had been vaccinated with bacillus of Calmette and Guerin (BCG), and none was HIV-1 positive. All parents of the children included in the study provided consent before the initiation of screening procedures. Chest radiography and Mantoux testing were carried out at the Pediatric Infectious Diseases Clinic, University of Sassari. The QFT microbiological examinations were done in the Mycobacteriology Laboratory, Department of Biomedical Science, University of Sassari. All of the children underwent the Mantoux skin test (Biocine Test PPD; Chiron, From the Department of Biomedical SciSiena, Italy); induration with a diameter ⬎ 5 mm at 48 to 72 hours was considered a ence (P.M., A.B., S.O., L.S., S.Z.) and Pedipositive result. The QFT results were scored as specified by the manufacturer (cutoff value atric Infectious Disease Clinic (G.M., P.O.), for a positive test, 0.35 IU/mL). University of Sassari, Sassari, Italy and Department of Public Health, Sassari, Italy Gastric aspirates were collected from children who were Mantoux- and/or QFT(R.D.). positive; staining and microscopy, nested polymerase chain reaction (PCR), and culture Submitted for publication Jul 5, 2007; last 5-7 tests were performed following standard protocols. In children with active TB, a DNA revision received Nov 6, 2007; accepted 8 Dec 5, 2007. fingerprinting assay was performed according to standard protocols. Reprint requests: Paola Molicotti, DepartStatistical analysis was elaborated using Fisher’s exact test. ment of Biomedical Sciences, University of Sassari, Viale San Pietro 43/b, 07100 Sassari, Italy. E-mail: [email protected].
BCG LTBI PCR
Bacillus of Calmette and Guerin Latent tuberculosis infection Polymerase chain reaction
QFT TB
QuantiFERON-TB Gold Tuberculosis
0022-3476/$ - see front matter Copyright © 2008 Mosby Inc. All rights reserved. 10.1016/j.jpeds.2007.12.014
585
Table. Summary of Mantoux test, chest radiography and QTF test findings Mantoux
Radiograph
QFT
19 10 29
11* 18† 29
21 8 29
Positive Negative Total
*Includes 5 children with primary complex and 6 children with hilar adenopathy. †One child had inconclusive radiography findings.
RESULTS Chest radiograph revealed primary tubercle complex in 5 children, hilar adenopathy in 6 children, inconclusive findings in 1 child, and negative findings in 17 children (Table). The Mantoux test was positive in 19 children and negative in 10; the QFT test was positive in 21 children and negative in 8 (Table). The first child diagnosed with active TB and the child with an inconclusive radiograph had a negative Mantoux test at first screening; repetition of the same test 2 weeks later yielded positive results. Both of these children had a positive QTF test at the first screening. All of the QFTnegative children underwent a repeat assay 2 months later, and all results were again negative. All staining tests performed on direct specimens were negative. Six children were culture-positive in both the MGIT 960 and Lowenstein-Jensen media; these 6 specimens also were positive for M tuberculosis on nested PCR assay. DNA fingerprinting analysis indicated that all children were infected by the same strain of M tuberculosis (data not shown). No statistically significant difference in results between the Mantoux and QFT tests (2-tailed P value ⫽ .77) was observed.
DISCUSSION Our results indicate that the QFT test is effective in identifying LTBI in children. Statistical analysis demonstrated no significant difference in accuracy between the Mantoux test and the QFT test. The delay of 3 months from the time of infection to Mantoux test conversion is problematic,9 because the time span between latent to active infection may be less than 3 months, especially in children. An alternative system that can provide earlier detection of TB is valuable for preventive treatment. In this outbreak, 2 children were QFTpositive but Mantoux-negative at first screening, and use of the QFT test allowed us to start the anti-TB treatment promptly. Another limitation of the Mantoux test is that a “booster effect” may occur after repeated use.10 In contrast, the QFT test can be performed repeatedly without affecting the M tuberculosis antigen–specific host immune response. This was very important in our study, because 8 children were both Mantoux- and QFT-negative. To exclude the presence of TB infection, the QFT test was performed repeatedly with
586
Molicotti et al
no fear of the booster reaction. After 2 months, the 8 children were QFT-negative, and thus LTBI was excluded. A very high percentage of the infected children developed TB. This may be due to the lack of BCG vaccination, high inoculum, or virulence of the transmitted M tuberculosis strain. The children with active TB were all QFT-positive, but their small number makes it impossible to evaluate the sensitivity of QFT in these patients. We observed that the first test may be more sensitive (albeit only slightly so), and that immediate treatment may be administered. Further studies are needed to demonstrate the usefulness of this assay in diagnosing active TB in children. Results of DNA fingerprinting indicated that infection from one child to another occurred within the same class, but the index case remained unclear. This is important because TB is usually transmitted by an adult, and after investigating school and family background, no case of active TB could be identified. One limitation of our study is the small number of children analyzed. Statistical analysis found no significant differences in accuracy between the QFT and Mantoux tests. Taking into consideration the QFT test’s higher specificity, possible earlier and greater sensitivity, lack of booster effect, and objective interpretation of the QFT test results compared with Mantoux, the QFT might be used initially in conjunction with, and eventually even be substituted for, the Mantoux test for diagnosing TB and LTBI in children.
REFERENCES 1. Shingadia D, Novelli V. Diagnosis and treatment of tuberculosis in children. Lancet Infect Dis 2003;3:624-32. 2. Pai M. Alternatives to the tuberculin skin test: interferon-gamma assays in the diagnosis of Mycobacterium tuberculosis infection. Indian J Med Microbiol 2005;23:151-8. 3. Mori T, Sakatani M, Yamagishi F, Takashima T, Kawabe Y, Nagao K, et al. Specific detection of tuberculosis infection: an interferon-gamma based assay using new antigens. Am J Respir Crit Care Med 2004;170:59-64. 4. Arend SM, Engelhard AC, Groot G, de Boer K, Andersen P, Ottenhoff TH, et al. Tuberculin skin testing compared with T-cell responses to Mycobacterium tuberculosis–specific and –nonspecific antigens for detection of latent infection in person with recent tuberculosis contact. Clin Diagn Lab Immunol 2001;8:1089-96. 5. Ardito F, Posteraro B, Sanguinetti M, Zanetti S, Fadda G. Evaluation of the BACTEC mycobacteria growth indicator tube (MGIT 960) automated system for drug susceptibility testing of Mycobacterium tuberculosis. J Clin Microbiol 2001;39:4440-4. 6. Ginesu F, Pirina P, Sechi LA, Molicotti P, Santoru L, Porcu L, et al. Microbiological diagnosis of tuberculosis: a comparison of old and new methods. J Chemother 1998;10:295-300. 7. American Thoracic Society. Diagnostic standards and classification of tuberculosis in adults and children. Am J Respir Crit Care Med 2000;161:1376-95. 8. Sechi LA, Zanetti S, Dupre I, Aceti A, Sanguinetti M, Fadda G. Genotypic changes in DNA fingerprinting patterns of Mycobacterium tuberculosis strains from HIV-positive persons in Sardinia. AIDS 1998;12:2084-6. 9. Starke JR. Pediatric tuberculosis: new time for a new approach. Tuberculosis 2003;83:208-12. 10. Mazurek GH, Jereb J, Lobue P, Iademarco MF, Metchock B, Vernon A, Division of Tuberculosis Elimination, National Center for HIV, STD, and TB Prevention, Centers for Disease Control and Prevention. Guidelines for using the QuantiFERON-TB gold test for detecting Mycobacterium tuberculosis infection, United States. MMWR Recomm Rep 2005; 54(RR-15):49-55. Erratum 54:1288.
The Journal of Pediatrics • April 2008
This study compared the effectiveness of the QuantiFERON-TB Gold (QFT) assay with the Mantoux tuberculin skin test to detect Mycobacterium tuberculosis infection in 29 children during a school outbreak of tuberculosis. Of the 21 children with M tuberculosis infection, 11 had a radiograph suggestive of the infection. The QFT assay was positive in all 21 of the children, and the Mantoux test was negative at first testing in 2 children (1 of whom was the sentinel case). The findings demonstrate that the QFT test is extremely useful in accurately identifying infected and uninfected children, permitting rapid intervention. (J Pediatr 2008;152:585-6)
iagnosis of tuberculosis (TB) in children is based on clinical examination, chest radiography, and the Mantoux test.1 Until recently, despite its limitations,2 the Mantoux test was the only tool available for the diagnosis of latent tuberculosis infection (LTBI). QuantiFERON-TB Gold (QFT) (Cellestis Limited, Carnegie, Victoria, Australia) is an in vitro immunologic assay that measures the interferon-gamma released by T lymphocytes sensitized with the Mycobacterium tuberculosis–specific antigens ESAT-6 and CFP-10. The specificity and sensitivity of this assay has been demonstrated in adults, and excellent results have been obtained for both active TB3 and LTBI.4 In early summer 2006, a 10-year-old child was admitted to the Pediatric Infectious Diseases Clinic of the University of Sassari with fever, cough, and a chest radiograph compatible with a primary TB complex. Subsequently, the Mantoux and QFT tests were performed. M tuberculosis was isolated in culture from the gastric aspirate, and the molecular test was positive for M tuberculosis as well. Immediate contact investigation (chest radiograph, Mantoux, QFT) was performed for all of the child’s fifth-grade classmates and for other children in different classrooms attending the same school. This outbreak of TB prompted us to investigate the effectiveness of the QFT assay and to compare the results with the Mantoux test as a tool for diagnosing TB in children.
D
METHODS A total of 29 children (16 males and 13 females age 6 to 10 years, 19 of whom were the sentinel case’s classmates), were examined. None of the children had been vaccinated with bacillus of Calmette and Guerin (BCG), and none was HIV-1 positive. All parents of the children included in the study provided consent before the initiation of screening procedures. Chest radiography and Mantoux testing were carried out at the Pediatric Infectious Diseases Clinic, University of Sassari. The QFT microbiological examinations were done in the Mycobacteriology Laboratory, Department of Biomedical Science, University of Sassari. All of the children underwent the Mantoux skin test (Biocine Test PPD; Chiron, From the Department of Biomedical SciSiena, Italy); induration with a diameter ⬎ 5 mm at 48 to 72 hours was considered a ence (P.M., A.B., S.O., L.S., S.Z.) and Pedipositive result. The QFT results were scored as specified by the manufacturer (cutoff value atric Infectious Disease Clinic (G.M., P.O.), for a positive test, 0.35 IU/mL). University of Sassari, Sassari, Italy and Department of Public Health, Sassari, Italy Gastric aspirates were collected from children who were Mantoux- and/or QFT(R.D.). positive; staining and microscopy, nested polymerase chain reaction (PCR), and culture Submitted for publication Jul 5, 2007; last 5-7 tests were performed following standard protocols. In children with active TB, a DNA revision received Nov 6, 2007; accepted 8 Dec 5, 2007. fingerprinting assay was performed according to standard protocols. Reprint requests: Paola Molicotti, DepartStatistical analysis was elaborated using Fisher’s exact test. ment of Biomedical Sciences, University of Sassari, Viale San Pietro 43/b, 07100 Sassari, Italy. E-mail: [email protected].
BCG LTBI PCR
Bacillus of Calmette and Guerin Latent tuberculosis infection Polymerase chain reaction
QFT TB
QuantiFERON-TB Gold Tuberculosis
0022-3476/$ - see front matter Copyright © 2008 Mosby Inc. All rights reserved. 10.1016/j.jpeds.2007.12.014
585
Table. Summary of Mantoux test, chest radiography and QTF test findings Mantoux
Radiograph
QFT
19 10 29
11* 18† 29
21 8 29
Positive Negative Total
*Includes 5 children with primary complex and 6 children with hilar adenopathy. †One child had inconclusive radiography findings.
RESULTS Chest radiograph revealed primary tubercle complex in 5 children, hilar adenopathy in 6 children, inconclusive findings in 1 child, and negative findings in 17 children (Table). The Mantoux test was positive in 19 children and negative in 10; the QFT test was positive in 21 children and negative in 8 (Table). The first child diagnosed with active TB and the child with an inconclusive radiograph had a negative Mantoux test at first screening; repetition of the same test 2 weeks later yielded positive results. Both of these children had a positive QTF test at the first screening. All of the QFTnegative children underwent a repeat assay 2 months later, and all results were again negative. All staining tests performed on direct specimens were negative. Six children were culture-positive in both the MGIT 960 and Lowenstein-Jensen media; these 6 specimens also were positive for M tuberculosis on nested PCR assay. DNA fingerprinting analysis indicated that all children were infected by the same strain of M tuberculosis (data not shown). No statistically significant difference in results between the Mantoux and QFT tests (2-tailed P value ⫽ .77) was observed.
DISCUSSION Our results indicate that the QFT test is effective in identifying LTBI in children. Statistical analysis demonstrated no significant difference in accuracy between the Mantoux test and the QFT test. The delay of 3 months from the time of infection to Mantoux test conversion is problematic,9 because the time span between latent to active infection may be less than 3 months, especially in children. An alternative system that can provide earlier detection of TB is valuable for preventive treatment. In this outbreak, 2 children were QFTpositive but Mantoux-negative at first screening, and use of the QFT test allowed us to start the anti-TB treatment promptly. Another limitation of the Mantoux test is that a “booster effect” may occur after repeated use.10 In contrast, the QFT test can be performed repeatedly without affecting the M tuberculosis antigen–specific host immune response. This was very important in our study, because 8 children were both Mantoux- and QFT-negative. To exclude the presence of TB infection, the QFT test was performed repeatedly with
586
Molicotti et al
no fear of the booster reaction. After 2 months, the 8 children were QFT-negative, and thus LTBI was excluded. A very high percentage of the infected children developed TB. This may be due to the lack of BCG vaccination, high inoculum, or virulence of the transmitted M tuberculosis strain. The children with active TB were all QFT-positive, but their small number makes it impossible to evaluate the sensitivity of QFT in these patients. We observed that the first test may be more sensitive (albeit only slightly so), and that immediate treatment may be administered. Further studies are needed to demonstrate the usefulness of this assay in diagnosing active TB in children. Results of DNA fingerprinting indicated that infection from one child to another occurred within the same class, but the index case remained unclear. This is important because TB is usually transmitted by an adult, and after investigating school and family background, no case of active TB could be identified. One limitation of our study is the small number of children analyzed. Statistical analysis found no significant differences in accuracy between the QFT and Mantoux tests. Taking into consideration the QFT test’s higher specificity, possible earlier and greater sensitivity, lack of booster effect, and objective interpretation of the QFT test results compared with Mantoux, the QFT might be used initially in conjunction with, and eventually even be substituted for, the Mantoux test for diagnosing TB and LTBI in children.
REFERENCES 1. Shingadia D, Novelli V. Diagnosis and treatment of tuberculosis in children. Lancet Infect Dis 2003;3:624-32. 2. Pai M. Alternatives to the tuberculin skin test: interferon-gamma assays in the diagnosis of Mycobacterium tuberculosis infection. Indian J Med Microbiol 2005;23:151-8. 3. Mori T, Sakatani M, Yamagishi F, Takashima T, Kawabe Y, Nagao K, et al. Specific detection of tuberculosis infection: an interferon-gamma based assay using new antigens. Am J Respir Crit Care Med 2004;170:59-64. 4. Arend SM, Engelhard AC, Groot G, de Boer K, Andersen P, Ottenhoff TH, et al. Tuberculin skin testing compared with T-cell responses to Mycobacterium tuberculosis–specific and –nonspecific antigens for detection of latent infection in person with recent tuberculosis contact. Clin Diagn Lab Immunol 2001;8:1089-96. 5. Ardito F, Posteraro B, Sanguinetti M, Zanetti S, Fadda G. Evaluation of the BACTEC mycobacteria growth indicator tube (MGIT 960) automated system for drug susceptibility testing of Mycobacterium tuberculosis. J Clin Microbiol 2001;39:4440-4. 6. Ginesu F, Pirina P, Sechi LA, Molicotti P, Santoru L, Porcu L, et al. Microbiological diagnosis of tuberculosis: a comparison of old and new methods. J Chemother 1998;10:295-300. 7. American Thoracic Society. Diagnostic standards and classification of tuberculosis in adults and children. Am J Respir Crit Care Med 2000;161:1376-95. 8. Sechi LA, Zanetti S, Dupre I, Aceti A, Sanguinetti M, Fadda G. Genotypic changes in DNA fingerprinting patterns of Mycobacterium tuberculosis strains from HIV-positive persons in Sardinia. AIDS 1998;12:2084-6. 9. Starke JR. Pediatric tuberculosis: new time for a new approach. Tuberculosis 2003;83:208-12. 10. Mazurek GH, Jereb J, Lobue P, Iademarco MF, Metchock B, Vernon A, Division of Tuberculosis Elimination, National Center for HIV, STD, and TB Prevention, Centers for Disease Control and Prevention. Guidelines for using the QuantiFERON-TB gold test for detecting Mycobacterium tuberculosis infection, United States. MMWR Recomm Rep 2005; 54(RR-15):49-55. Erratum 54:1288.
The Journal of Pediatrics • April 2008