Both assays rest on the same biological foundation: the cell-mediated immune response that T lymphocytes mount against specific Mycobacterium tuberculosis antigens. When sensitized T cells encounter peptides from the TB-specific proteins ESAT-6 and CFP-10, they release interferon-gamma, a signaling molecule whose presence reveals antigen-specific immune memory. QuantiFERON-TB Gold Plus, manufactured by Qiagen, incubates whole venous blood overnight with these antigens and then measures the accumulated IFNγ using an enzyme-linked immunosorbent assay, or ELISA. T-SPOT.TB, from Oxford Immunotec, takes a different route: it first isolates peripheral blood mononuclear cells from the blood, standardizes their number, and then counts the individual IFNγ-secreting T cells with an enzyme-linked immunospot assay, known as ELISPOT. QuantiFERON additionally includes peptides recognized by TB-specific CD8+ T lymphocytes, broadening the immune cell populations it stimulates beyond the CD4+ compartment.

The study, conducted at the Ramus laboratory between January 1 and December 31, 2024, processed a total of 7,392 patient samples: 6,887 with QuantiFERON-TB Gold Plus and 505 with T-SPOT.TB. The outcome distributions differed markedly between the two platforms. QuantiFERON yielded negative results in an expected 92.3 percent of samples and positive results in only 5.3 percent, reflecting the low TB prevalence in Bulgaria, a high-income country with established tuberculosis control programs. T-SPOT.TB, by contrast, produced 59.8 percent negative and 24.6 percent positive results, a substantially higher positivity rate that hints at its greater analytical sensitivity in populations with low T-cell counts, such as immunosuppressed patients.

Perhaps the most striking differences emerged in the gray zones of testing. Borderline and indeterminate results, the categories that most frustrate clinicians, hovered around 2 percent for QuantiFERON but reached 5.7 percent and 9.5 percent, respectively, for T-SPOT.TB. The indeterminate rate for T-SPOT.TB dropped to 5.3 percent when 21 patients on biological therapy were followed up and their results subsequently converted to negative. QuantiFERON recorded indeterminate results in only 0.4 percent of cases overall, rising to 1.4 percent among patients receiving biological therapy, while 0.5 percent of its results were classified as borderline and 0.1 percent produced no valid outcome. T-SPOT.TB, meanwhile, classified 5.7 percent of results as borderline and failed to generate a valid result in 0.4 percent of tests.

To probe the clinical meaning of these differences, the researchers analyzed 94 patients who underwent both tests, either simultaneously or within defined intervals of up to one year, along with 12 patients who were tested multiple times with a single method. The dual-tested cohort had a mean age of 52 years, ranging from 21 to 85, and comprised 44 females and 50 males. Concordance between the two assays appeared in 51 individuals, or 54.3 percent: 31 had concordant positive results, 17 had concordant negative results, and 3 showed indeterminate results in both tests. Discrepancies surfaced in the remaining 43 patients, or 45.7 percent, a proportion that demands careful clinical interpretation.

The analysis of these discordant cases revealed a reassuring pattern. In 61 percent of the discrepancies, or 26 of 43 patients, results shifted from indeterminate or borderline to negative, or the reverse, within one to six months, a degree of variability the authors deem acceptable under existing guidelines. Among the 17 individuals, or 39 percent, whose results changed from positive to negative or indeterminate, further investigation showed that all had initiated biological therapy. This finding carries significant weight: immunosuppressive treatments can suppress the T-cell response that both assays measure, converting a previously detectable antigen-specific reaction into an undetectable one. Among the 12 patients repeatedly tested with T-SPOT.TB over intervals of four to eight months, three showed consistent positive results, two showed consistent negative results, and seven experienced changes, all of whom were on biological therapy.

Statistical analysis quantified the agreement between the two platforms. Cohen’s kappa, a measure of inter-rater reliability, came in at 0.312, indicating significant but fair agreement, far from the perfect concordance of 1. However, when indeterminate and borderline results were omitted from the calculation, the agreement became substantial, with a kappa of 0.697, much closer to perfect. Under this refined analysis, 85.7 percent of samples were concordant while 14.3 percent were discordant. Sensitivity and specificity also diverged depending on which test served as the reference: T-SPOT.TB, compared to QuantiFERON, showed a sensitivity of 83.78 percent and a specificity of 89.47 percent, while QuantiFERON, compared to T-SPOT.TB, demonstrated a sensitivity of 93.94 percent and a specificity of 73.91 percent.

Each assay carries distinct methodological advantages that shape its clinical niche. T-SPOT.TB, as an ELISPOT-based assay, standardizes the number of peripheral blood mononuclear cells, which may provide greater sensitivity in immunocompromised patients or those with lymphopenia, since it reduces the impact of low cell counts on the result. QuantiFERON, as a whole-blood ELISA-based assay, is technically simpler, less labor-intensive, and more easily standardized, making it better suited for high-throughput laboratory settings. Its inclusion of antigen tubes designed to stimulate both CD4+ and CD8+ T-cell responses may offer advantages in certain clinical contexts. The choice between them, the authors suggest, should depend on patient characteristics, laboratory capacity, and clinical indication rather than on any universal superiority of one platform.

The study also spotlighted a compelling clinical case: a 55-year-old man with a persistent subfebrile condition and swelling in the area of one testicle, who consulted a urologist and underwent periodic antibiotic therapy with isoniazid without lasting effect. Roughly a decade earlier, he had tested positive by QuantiFERON. When retested twice within about a month, both QuantiFERON and T-SPOT.TB returned positive results, pointing to extrapulmonary tuberculosis after a pulmonary form of the disease was ruled out. Six months of specific anti-TB therapy produced a lasting effect on his clinical symptoms, and he has remained stable since, with no chills, fatigue, or fever. A specialist in extrapulmonary tuberculosis indicated that the test may remain positive for a prolonged period, illustrating how IGRA positivity can persist even after successful treatment.

The broader context reinforces the value of these findings. A health technology assessment by Ontario Health, encompassing 12 systematic reviews and more than 500 primary studies, demonstrated that IGRAs serve as effective rule-in tests for latent tuberculosis infection due to consistently high specificity, outperforming the tuberculin skin test in reducing false positives, particularly among immunocompromised populations, children, elderly individuals, and BCG-vaccinated persons. Economic analyses from five Canadian studies revealed that IGRA testing proved cost-effective or cost-saving for high-risk latent TB populations, with healthcare providers expressing strong preference for the single-visit requirement compared to the multiple visits demanded by skin testing. Yet the ability of IGRAs to yield indeterminate results in immunocompromised patients signals the need for further clinical investigation when skin tests may produce false negatives.

Both assays share inherent limitations. Neither can reliably differentiate between latent TB infection and active TB disease, and both depend on a robust host immune response, meaning false negatives may occur in immunocompromised individuals, young children, or patients with advanced disease. A recent systematic review and meta-analysis of IGRAs in people living with HIV, covering 6,525 patients from 45 studies, found that while both tests demonstrated acceptable specificity, their sensitivity was lower, particularly for latent TB infection, suggesting IGRAs may be more reliable for detecting active TB than for ruling out latent infection in immunocompromised populations. The variation in antigen-specific T-cell responses observed in the current cohort may also reflect differences in exposure history, BCG vaccination status, or concurrent infections.

From a clinical perspective, discrepancies between QuantiFERON and T-SPOT.TB results necessitate careful interpretation in conjunction with epidemiological risk factors, radiological findings, and clinical presentation. For borderline or discordant cases, repeating the IGRA or performing both tests sequentially may improve diagnostic confidence, particularly in high-risk groups. The retrospective nature of the study and the lack of detailed clinical data on individual disease activity, concomitant medications, and duration of biological therapy represent limitations that future prospective designs should address through detailed patient stratification and combined analysis of qualitative and quantitative results.

The bottom line for clinicians is clear: the results from both IFNγ-based tests are comparable, with acceptable variation over time, and either method can be relied upon in routine laboratory diagnostics. However, significant changes in results, whether a sharp conversion from positive to negative or the emergence of indeterminate readings, should always be interpreted in light of ongoing specific or biological therapy. As tuberculosis control efforts intensify worldwide and biological therapies continue to expand across medicine, this real-world evidence offers laboratories and physicians a practical roadmap for navigating the nuances of interferon-gamma testing in the patients who need it most.

Subject of Research: Comparative diagnostic performance of two interferon-gamma release assays for Mycobacterium tuberculosis infection

Article Title: Comparative results of two IGRA tests for determining the specific response against Mycobacterium tuberculosis infection

Article References: Stoilov, P., Mihova, A., Mihaylov, R., Zlateva, S., Dimcheva, P., Pentchev, E., Kalcheva, M., Pencheva, B., Naseva, E., & Velikova, T. (2026). Comparative results of two IGRA tests for determining the specific response against Mycobacterium tuberculosis infection. Pharmacia, 73, Article e167649. https://doi.org/10.3897/pharmacia.73.e167649

Image Credits: AI Generated

DOI: 10.3897/pharmacia.73.e167649

Keywords: tuberculosis, IGRA, QuantiFERON, T-SPOT.TB, interferon-gamma, diagnostics, latent TB infection, biological therapy, Comparative, results, tests, determining

The research, published in PLOS Global Public Health, was led by Hannah M. Rickman of the Malawi-Liverpool-Wellcome Trust clinical research programme together with collaborators from the Malawi Ministry of Health, the London School of Hygiene & Tropical Medicine and the University of Liverpool. The team set out to answer a deceptively simple question: where in a large African city is Mycobacterium tuberculosis, the bacterium that causes tuberculosis, actively being transmitted right now? Their answer came not from counting sick adults, but from measuring silent infection in healthy toddlers.

The logic behind the approach rests on a fundamental feature of TB epidemiology. Young children who test positive for immune reactivity against M. tuberculosis must have been infected recently, because the window between birth and testing is short. Unlike adults, who may carry latent infection acquired decades earlier, children aged one to four years serve as sensitive biological sensors of ongoing transmission in their immediate surroundings. Every positive result in a toddler points to an undiagnosed infectious case somewhere nearby, most likely an adult in the household or neighbourhood whose illness has never reached a clinic.

To exploit this signal, the researchers conducted a cross-sectional survey across 33 high-burden neighbourhoods of Blantyre, a city of roughly one million people in southern Malawi. They enrolled 1,545 healthy children between one and four years of age using two complementary recruitment strategies. Some families were approached through health facilities, where children attend for routine immunisations and growth monitoring, while others were identified through random household sampling guided by satellite imagery of the urban landscape. This hybrid design was deliberately pragmatic, designed to be affordable and repeatable by TB programmes in resource-limited settings rather than requiring the expensive logistics of a classic disease prevalence survey.

Each child provided a small blood sample that was tested with an interferon-gamma release assay, or IGRA, a laboratory test that detects the immune response of T cells to proteins specific to M. tuberculosis. Unlike tuberculin skin tests, IGRAs are not confounded by prior vaccination with bacille Calmette-Guérin, the live vaccine given at birth across most of sub-Saharan Africa. A positive QFT-Plus result therefore indicates genuine immunological exposure to the tuberculosis bacterium, whether the child has cleared the infection, is carrying it latently, or in rare cases has progressed to active disease.

The headline finding was sobering. Of the 1,545 children tested, 102, or 6.6 percent, had a positive IGRA result. Because infection accumulates over the first years of life, the researchers converted this prevalence into an annual risk of M. tuberculosis infection of 2.7 percent, with a 95 percent credible interval of 2.2 to 3.2 percent. In practical terms, roughly one in every 37 children in these neighbourhoods acquires a new tuberculosis infection each year, a rate that speaks to a level of ongoing community transmission far higher than most residents or health workers would suspect from clinic-based figures alone.

The social gradient in the results was equally telling. Children living in the poorest third of households were substantially more likely to be immunoreactive than those in the richest third, with 8.7 percent positivity compared with 4.9 percent. After adjusting for other factors in a hierarchical Bayesian logistic regression model that accounted for clustering within neighbourhoods, the poorest children had nearly double the odds of a positive result, with an adjusted odds ratio of 1.88. Notably, the risk was not explained by the usual suspects of paediatric TB vulnerability: exposure to HIV, malnutrition, and reported contact with a household member with TB all showed no significant association with immunoreactivity in this analysis.

Perhaps the most consequential finding concerned geography. The researchers observed substantial heterogeneity between neighbourhoods, with estimated annual risks of infection ranging from 1.1 percent to 4.1 percent across the 33 communities studied. Yet when they compared these neighbourhood-level infection risks against official TB case notification rates, the correlation was absent. Areas reporting the most tuberculosis cases to health authorities were not the areas where children were most likely to be acquiring infection. The authors attribute this disconnect to disparities in healthcare access: case notifications reflect who reaches a clinic and gets diagnosed, not who is actually transmitting the disease. In a city where diagnostic capacity, transport costs and health-seeking behaviour vary sharply from one district to the next, notification data can point programmes toward the wrong neighbourhoods entirely.

The statistical machinery behind these conclusions was designed to handle sparse data and clustered sampling. The hierarchical Bayesian logistic regression framework allowed the team to estimate infection risk at both the individual and neighbourhood levels simultaneously, borrowing strength across communities while quantifying uncertainty through credible intervals rather than single point estimates. Neighbourhood-level random effects captured the unexplained variation between communities, and the model structure made it possible to test whether household wealth, HIV exposure, nutritional status, reported TB contact or notification rates predicted positivity after accounting for this spatial clustering. This approach, the authors argue, is what makes the survey a template for surveillance rather than a one-off research exercise.

The implications reach well beyond Blantyre. Tuberculosis prevalence surveys, in which teams screen entire populations for active disease, were once the gold standard for measuring epidemic trends, but they have become prohibitively expensive as countries approach elimination targets and disease becomes rare. Passive surveillance through case notifications, meanwhile, is systematically biased by unequal access to care. Measuring immunoreactivity in young children offers a third way: an indirect but biologically grounded readout of recent transmission that requires only a blood test, a laboratory and a sampling frame. The Blantyre study demonstrates that such surveys can be run at scale using convenience recruitment in clinics supplemented by satellite-guided household sampling, keeping costs within reach of national TB programmes.

For the children of Blantyre, the findings carry an immediate message. The burden of recent tuberculosis infection is high, concentrated among the poorest families, and invisible to the surveillance systems currently guiding resource allocation. The authors argue that M. tuberculosis immunoreactivity surveys could help programmes identify high-risk populations, monitor trends over time and target interventions such as active case finding and preventive therapy to the neighbourhoods where transmission actually occurs. In a global campaign that has pledged to end the tuberculosis epidemic, knowing precisely where the epidemic lives may prove to be the most valuable diagnostic of all, and it may be found in a few millilitres of blood from a healthy two-year-old.

Subject of Research: Community-based interferon-gamma release assay survey measuring Mycobacterium tuberculosis immunoreactivity and transmission risk in young children in Blantyre, Malawi

Article Title: Heterogeneity in Mycobacterium tuberculosis immunoreactivity in young children in Blantyre, Malawi: A community-based survey

Article References: Rickman, H. M., Phiri, M. D., Mbale, H., Feasey, H. R. A., Nliwasa, M., Chagaluka, G., Seddon, J. A., Mwandumba, H. C., Horton, K. C., Henrion, M. Y. R., Mwenyenkulu, T., Mbendera, K. N., Nightingale, E. S., Corbett, E. L., & MacPherson, P. (2026). Heterogeneity in Mycobacterium tuberculosis immunoreactivity in young children in Blantyre, Malawi: A community-based survey. PLOS Global Public Health, 6(10), e0007269. https://doi.org/10.1371/journal.pgph.0007269

Image Credits: AI Generated

DOI: 10.1371/journal.pgph.0007269

Keywords: tuberculosis, Mycobacterium tuberculosis, IGRA, Malawi, Blantyre, annual risk of infection, children, surveillance, health inequity, Bayesian modelling, public health, epidemiology

The scale of the analysis is what sets it apart. Most previous research on migrant tuberculosis screening has focused narrowly on detecting active disease or on modelling the cost-effectiveness of screening for tuberculosis infection, the asymptomatic stage in which the bacteria persist without causing illness. Few studies have examined large-scale national programmes, reported age-stratified yields, or traced the full cascade of preventive treatment from diagnosis through completion. By linking screening results collected by all 25 Municipal Public Health Services to national notification data, the researchers were able to follow nearly 150,000 individuals from the moment of their first chest X-ray or infection test through to treatment outcomes.

The headline finding concerns the stark difference between two migrant categories. Among 87,511 immigrants, defined as labour migrants, international students and others arriving through regular channels, entry screening detected 69 cases of active tuberculosis, a yield of 79 cases per 100,000 people screened. Among 61,703 asylum seekers, screening identified 170 cases, a yield of 276 per 100,000, more than three times higher. The gap reflects both the higher tuberculosis burden in the countries from which asylum seekers predominantly arrive, including Eritrea, Afghanistan, Somalia and Algeria, and the adverse conditions of the migration journey itself, which can reactivate dormant infection through malnutrition, stress and crowding.

Country-level yields sharpened the picture further. Among asylum seekers, screening prevalence exceeded World Health Organization incidence estimates for their countries of birth in every case except Afghanistan, with the highest yields recorded among people from Sierra Leone at 888 per 100,000, Eritrea at 704, Somalia at 581, the Gambia at 562 and South Sudan and Sudan at 519. Among immigrants, by contrast, prevalence at entry was lower than WHO estimates for every country contributing more than five detected cases, with the highest yields among those from Indonesia at 291 per 100,000 and the Philippines at 241, and comparatively low yields among immigrants from India and South Africa. The researchers interpret this as evidence of the healthy migrant effect: people who migrate for work or study tend to be younger, healthier and more affluent than the general population of their country of origin, and therefore carry a lower tuberculosis risk than WHO national averages imply.

Perhaps the most consequential finding for clinical practice is the weakness of symptom-based detection. Of the 202 migrants diagnosed with pulmonary tuberculosis through screening, only 27.2 percent reported a cough, the symptom that anchors most passive case-finding strategies. The proportion was lower still among immigrants, at 13.8 percent, compared with 32.6 percent among asylum seekers. In other words, nearly three in four screen-detected pulmonary cases would have been missed by symptom screening alone. This aligns with a growing body of evidence on what the WHO now terms asymptomatic tuberculosis, and with community prevalence surveys in high-incidence settings showing that roughly half of bacteriologically confirmed pulmonary cases report no suggestive symptoms. Because such individuals can transmit the infection unknowingly for months, the study argues that systematic radiological screening retains clear value for populations at elevated risk, catching disease before symptom onset and before onward transmission occurs.

The diagnostic profile of detected cases supports the programme’s technical rigour. Overall, 56.5 percent of the 239 screen-detected tuberculosis cases were bacteriologically confirmed, rising to 63.9 percent among pulmonary cases, and 44 individuals had acid-fast bacilli visible on microscopy, indicating potentially infectious disease. Only two cases involved multidrug-resistant tuberculosis. Treatment outcomes, however, revealed a care gap: 92.8 percent of immigrants with tuberculosis were recorded as cured or having completed treatment, compared with 81.2 percent of asylum seekers, a difference driven mainly by loss to follow-up, including five asylum seekers who never initiated treatment and others who left the country before completing therapy. The authors point to the need for tailored support and stronger cross-border collaboration to maintain continuity of care for mobile populations.

The study’s second major contribution is its analysis of tuberculosis infection screening among more than 20,000 immigrants, the first time this component has been included in the Dutch national evaluations. Infection prevalence rose sharply with age: 0.6 percent among children under 12, 4.4 percent among adolescents aged 12 to 17, and 12.9 percent among adults screened from the highest-incidence countries. Among adults, prevalence exceeded 20 percent for immigrants from Kenya, Nigeria and Zimbabwe, while remaining markedly lower at 4.9 percent for those from South Africa. The Dutch algorithm, which uses an initial tuberculin skin test followed by confirmatory interferon gamma release assay when the skin test measures 10 millimetres or more, likely suppressed false positives; only about half of adults with a positive skin test had infection confirmed by IGRA, and confirmation rates fell below 10 percent in young children, where prior BCG vaccination may drive false-positive skin reactions.

The cascade of preventive treatment offered encouraging evidence that a well-structured public health system can convert infection diagnoses into completed therapy. Preventive treatment was initiated in 68.6 percent of infected children under 12, 85.2 percent of adolescents and 70.3 percent of adults, and among those who started, completion exceeded 92 percent in every age group, most commonly with a short three-month regimen of rifampicin and isoniazid. Combining initiation and completion, between 64.6 and 81.5 percent of infected individuals across age groups successfully finished preventive therapy, figures that outperform pooled international estimates of roughly 52 percent. The weaker uptake among young children, attributed largely to parental reluctance to medicate healthy children, combined with their very low infection prevalence, prompted the National TB Guideline Committee to recommend exempting immigrant children under 12 from mandatory screening, a concrete example of epidemiological data reshaping national policy.

The findings arrive at a moment when tuberculosis control in Europe increasingly depends on migrant-focused strategies. In 2024, 37.6 percent of people notified with tuberculosis in the EU and European Economic Area were of foreign origin, a share exceeding 80 percent in several countries including the Netherlands, which recorded just 4.3 cases per 100,000 population overall. The study supports maintaining divergent screening thresholds, with asylum seekers screened from countries with incidence of 50 per 100,000 or above and immigrants from countries at 100 or above, and it strengthens the economic case for infection-based follow-up screening, which Dutch modelling suggests is more cost-effective than repeated chest X-rays for asylum seekers. Emerging tuberculosis-antigen skin tests may further improve specificity and cost-effectiveness. The authors conclude that targeted screening of migrants, combining disease detection with infection testing and high-quality preventive treatment, is a core and demonstrably effective component of tuberculosis elimination in low-incidence settings, provided that screening strategies continue to be refined with timely, age-specific and group-specific national data.

Subject of Research: National evaluation of tuberculosis disease and tuberculosis infection screening among migrants in the Netherlands from 2019 to 2023

Article Title: Tuberculosis disease and infection screening among migrants in the Netherlands 2019–2023: a national retrospective observational study

Article References: de Vries, G., Mulder, A., Spruijt, I., Huisman, E., Schimmel, H., Siebers, M., Vrubleuskaya, N., Zandbelt-Smits, M., van Gageldonk-Lafeber, R., & van den Hof, S. (2026). Tuberculosis disease and infection screening among migrants in the Netherlands 2019–2023: a national retrospective observational study. The Lancet Regional Health – Europe, 70, Article 101856. https://doi.org/10.1016/j.lanepe.2026.101856

Image Credits: AI Generated

DOI: 10.1016/j.lanepe.2026.101856

Keywords: tuberculosis, migrant health, screening, asylum seekers, tuberculosis infection, preventive treatment, public health, Netherlands, epidemiology, low-incidence countries, chest X-ray, IGRA