The Union and Médecins Sans Frontières approach to operational research.

Operational research (OR) has become a hot topic at national meetings, international conferences and donor fora. The International Union Against Tuberculosis and Lung Disease (The Union) and Médecins Sans Frontières (MSF) Operational Centre Brussels strongly promote and implement OR with colleagues in low- and middle-income countries. Here we describe how the two organisations define OR, and explain the guiding principles and methodology that underpin the strategy for developing and expanding OR in those countries. We articulate The Union's and MSF's approach to supporting OR, highlighting the main synergies and differences. Then, using the Malawi National Tuberculosis Control Programme as an example, we show how OR can be embedded within tuberculosis control activities, leading to changes in policy and practice at the national level. We discuss the difficult, yet vitally important, issue of capacity building, and share our vision of a new paradigm of product-related training and performance-based OR fellowships as two ways of developing the necessary skills at country level to ensure research is actually performed. Finally, we highlight the need to consider and incorporate into practice the ethical components of OR. This is a key moment to be involved in OR. We are confident that in partnership with interested stakeholders, including the World Health Organization, we can stimulate the implementation of quality, relevant OR as an integral part of health service delivery that in turn will lead to better health for people, particularly for those living in the poorer parts of the world

A novel approach in the treatment of neuroendocrine gastrointestinal tumors: Additive antiproliferative effects of interferon-γ and meta-iodobenzylguanidine

BACKGROUND: Therapeutic options to effectively inhibit growth and spread of neuroendocrine gastrointestinal tumors are still limited. As both meta-iodobenzylguanidine (MIBG) and interferon-γ (IFNγ) cause antineoplastic effects in neuroendocrine gastrointestinal tumor cells, we investigated the antiproliferative effects of the combination of IFNγ and non-radiolabeled MIBG in neuroendocrine gut STC-1 and pancreatic carcinoid BON tumor cells. METHODS AND RESULTS: IFNγ receptors were expressed in both models. IFNγ dose- and time-dependently inhibited the growth of both STC-1 and of BON tumor cells with IC(50)-values of 95 ± 15 U/ml and 135 ± 10 U/ml, respectively. Above 10 U/ml IFNγ induced apoptosis-specific caspase-3 activity in a time-dependent manner in either cell line and caused a dose-dependent arrest in the S-phase of the cell cycle. Furthermore, IFNγ induced cytotoxic effects in NE tumor cells. The NE tumor-targeted drug MIBG is selectively taken up via norepinephrine transporters, thereby specifically inhibiting growth in NE tumor cells. Intriguingly, IFNγ treatment induced an upregulation of norepinephrine transporter expression in neuroendocrine tumors cells, as determined by semi-quantitative RT-PCR. Co-application of sub-IC(50 )concentrations of IFNγ and MIBG led to additive growth inhibitory effects, which were mainly due to increased cytotoxicity and S-phase arrest of the cell cycle. CONCLUSION: Our data show that IFNγ exerts antiproliferative effects on neuroendocrine gastrointestinal tumor cells by inducing cell cycle arrest, apoptosis and cytotoxicity. The combination of IFNγ with the NE tumor-targeted agent MIBG leads to effective growth control at reduced doses of either drug. Thus, the administration of IFNγ alone and more so, in combination with MIBG, is a promising novel approach in the treatment of neuroendocrine gastrointestinal tumors

Within-Subject Variability of Interferon-g Assay Results for Tuberculosis and Boosting Effect of Tuberculin Skin Testing: A Systematic Review

Background: Variability in interferon-gamma release assays (IGRAs) results for tuberculosis has implications for interpretation of results close to the cut-point, and for defining thresholds for test conversion and reversion. However, little is known about the within-subject variability (reproducibility) of IGRAs. Several national guidelines recommend a twostep testing procedure (tuberculin skin test [TST] followed by IGRA) for the diagnosis of LTBI. However, the effect of a preceding TST on subsequent IGRA results has been reported in studies with apparently conflicting results. Methodology/Findings: We conducted a systematic review to synthesize evidence on within-subject variability of IGRA results and the potential boosting effect of TST. We searched several databases and reviewed citations of previous reviews on IGRAs. We included studies using commercial IGRAs, in addition to non-commercial versions of the ELISPOT assay. Four studies, fulfilling our predefined criteria, examined within-subject variability and 13 studies evaluated TST effects on subsequent IGRA responses. Meta-analysis was not considered appropriate because of heterogeneity in study methods, assays, and populations. Although based on limited data, within-subject variability was present in all studies but the magnitude varied (16-80%) across studies. A TST induced ‘‘boosting’ ’ of IGRA responses was demonstrated in several studies and although more pronounced in IGRA-positive (i.e. sensitized) individuals, also occurred in a smaller but not insignificant proportion of IGRA-negative subjects. The TST appeared to affect IGRA responses only after 3 days and may apparentl

Xpert MTB/RIF Ultra assay for tuberculosis disease and rifampicin resistance in children

Background Every year, an estimated one million children and young adolescents become ill with tuberculosis, and around 226,000 of those children die. Xpert MTB/RIF Ultra (Xpert Ultra) is a molecular World Health Organization (WHO)‐recommended rapid diagnostic test that simultaneously detects Mycobacterium tuberculosis complex and rifampicin resistance. We previously published a Cochrane Review 'Xpert MTB/RIF and Xpert MTB/RIF Ultra assays for tuberculosis disease and rifampicin resistance in children'. The current review updates evidence on the diagnostic accuracy of Xpert Ultra in children presumed to have tuberculosis disease. Parts of this review update informed the 2022 WHO updated guidance on management of tuberculosis in children and adolescents. Objectives To assess the diagnostic accuracy of Xpert Ultra for detecting: pulmonary tuberculosis, tuberculous meningitis, lymph node tuberculosis, and rifampicin resistance, in children with presumed tuberculosis. Secondary objectives To investigate potential sources of heterogeneity in accuracy estimates. For detection of tuberculosis, we considered age, comorbidity (HIV, severe pneumonia, and severe malnutrition), and specimen type as potential sources. To summarize the frequency of Xpert Ultra trace results. Search methods We searched the Cochrane Infectious Diseases Group Specialized Register, MEDLINE, Embase, three other databases, and three trial registers without language restrictions to 9 March 2021. Selection criteria Cross‐sectional and cohort studies and randomized trials that evaluated Xpert Ultra in HIV‐positive and HIV‐negative children under 15 years of age. We included ongoing studies that helped us address the review objectives. We included studies evaluating sputum, gastric, stool, or nasopharyngeal specimens (pulmonary tuberculosis), cerebrospinal fluid (tuberculous meningitis), and fine needle aspirate or surgical biopsy tissue (lymph node tuberculosis). For detecting tuberculosis, reference standards were microbiological (culture) or composite reference standard; for stool, we also included Xpert Ultra performed on a routine respiratory specimen. For detecting rifampicin resistance, reference standards were drug susceptibility testing or MTBDRplus. Data collection and analysis Two review authors independently extracted data and, using QUADAS‐2, assessed methodological quality judging risk of bias separately for each target condition and reference standard. For each target condition, we used the bivariate model to estimate summary sensitivity and specificity with 95% confidence intervals (CIs). We stratified all analyses by type of reference standard. We summarized the frequency of Xpert Ultra trace results; trace represents detection of a very low quantity of Mycobacterium tuberculosis DNA. We assessed certainty of evidence using GRADE. Main results We identified 14 studies (11 new studies since the previous review). For detection of pulmonary tuberculosis, 335 data sets (25,937 participants) were available for analysis. We did not identify any studies that evaluated Xpert Ultra accuracy for tuberculous meningitis or lymph node tuberculosis. Three studies evaluated Xpert Ultra for detection of rifampicin resistance. Ten studies (71%) took place in countries with a high tuberculosis burden based on WHO classification. Overall, risk of bias was low. Detection of pulmonary tuberculosis Sputum, 5 studies Xpert Ultra summary sensitivity verified by culture was 75.3% (95% CI 64.3 to 83.8; 127 participants; high‐certainty evidence), and specificity was 97.1% (95% CI 94.7 to 98.5; 1054 participants; high‐certainty evidence). Gastric aspirate, 7 studies Xpert Ultra summary sensitivity verified by culture was 70.4% (95% CI 53.9 to 82.9; 120 participants; moderate‐certainty evidence), and specificity was 94.1% (95% CI 84.8 to 97.8; 870 participants; moderate‐certainty evidence). Stool, 6 studies Xpert Ultra summary sensitivity verified by culture was 56.1% (95% CI 39.1 to 71.7; 200 participants; moderate‐certainty evidence), and specificity was 98.0% (95% CI 93.3 to 99.4; 1232 participants; high certainty‐evidence). Nasopharyngeal aspirate, 4 studies Xpert Ultra summary sensitivity verified by culture was 43.7% (95% CI 26.7 to 62.2; 46 participants; very low‐certainty evidence), and specificity was 97.5% (95% CI 93.6 to 99.0; 489 participants; high‐certainty evidence). Xpert Ultra sensitivity was lower against a composite than a culture reference standard for all specimen types other than nasopharyngeal aspirate, while specificity was similar against both reference standards. Interpretation of results In theory, for a population of 1000 children: • where 100 have pulmonary tuberculosis in sputum (by culture): ‐ 101 would be Xpert Ultra‐positive, and of these, 26 (26%) would not have pulmonary tuberculosis (false positive); and ‐ 899 would be Xpert Ultra‐negative, and of these, 25 (3%) would have tuberculosis (false negative). • where 100 have pulmonary tuberculosis in gastric aspirate (by culture): ‐ 123 would be Xpert Ultra‐positive, and of these, 53 (43%) would not have pulmonary tuberculosis (false positive); and ‐ 877 would be Xpert Ultra‐negative, and of these, 30 (3%) would have tuberculosis (false negative). • where 100 have pulmonary tuberculosis in stool (by culture): ‐ 74 would be Xpert Ultra‐positive, and of these, 18 (24%) would not have pulmonary tuberculosis (false positive); and ‐ 926 would be Xpert Ultra‐negative, and of these, 44 (5%) would have tuberculosis (false negative). • where 100 have pulmonary tuberculosis in nasopharyngeal aspirate (by culture): ‐ 66 would be Xpert Ultra‐positive, and of these, 22 (33%) would not have pulmonary tuberculosis (false positive); and ‐ 934 would be Xpert Ultra‐negative, and of these, 56 (6%) would have tuberculosis (false negative). Detection of rifampicin resistance Xpert Ultra sensitivity was 100% (3 studies, 3 participants; very low‐certainty evidence), and specificity range was 97% to 100% (3 studies, 128 participants; low‐certainty evidence). Trace results Xpert Ultra trace results, regarded as positive in children by WHO standards, were common. Xpert Ultra specificity remained high in children, despite the frequency of trace results. Authors' conclusions We found Xpert Ultra sensitivity to vary by specimen type, with sputum having the highest sensitivity, followed by gastric aspirate and stool. Nasopharyngeal aspirate had the lowest sensitivity. Xpert Ultra specificity was high against both microbiological and composite reference standards. However, the evidence base is still limited, and findings may be imprecise and vary by study setting. Although we found Xpert Ultra accurate for detection of rifampicin resistance, results were based on a very small number of studies that included only three children with rifampicin resistance. Therefore, findings should be interpreted with caution. Our findings provide support for the use of Xpert Ultra as an initial rapid molecular diagnostic in children being evaluated for tuberculosis

A Three-Way Comparison of Tuberculin Skin Testing, QuantiFERON-TB Gold and T-SPOT.TB in Children

BACKGROUND: There are limited data comparing the performance of the two commercially available interferon gamma (IFN-gamma) release assays (IGRAs) for the diagnosis of tuberculosis (TB) in children. We compared QuantiFERON-TB gold In Tube (QFT-IT), T-SPOT.TB and the tuberculin skin test (TST) in children at risk for latent TB infection or TB disease. METHODS AND FINDINGS: The results of both IGRAs were compared with diagnosis assigned by TST-based criteria and assessed in relation to TB contact history. Results from the TST and at least one assay were available for 96 of 100 children. Agreement between QFT-IT and T-SPOT.TB was high (93% agreement, kappa = 0.83). QFT-IT and T-SPOT.TB tests were positive in 8 (89%) and 9 (100%) children with suspected active TB disease. There was moderate agreement between TST and either QFT-IT (75%, kappa = 0.50) or T-SPOT.TB (75%, kappa = 0.51). Among 38 children with TST-defined latent TB infection, QFT-IT gold and T-SPOT.TB assays were positive in 47% and 39% respectively. Three TST-negative children were positive by at least one IGRA. Children with a TB contact were more likely than children without a TB contact to have a positive IGRA (QFT-IT LR 3.9; T-SPOT.TB LR 3.9) and a positive TST (LR 1.4). Multivariate linear regression analysis showed that the magnitude of both TST induration and IGRA IFN-gamma responses was significantly influenced by TB contact history, but only the TST was influenced by age. CONCLUSIONS: Although a high level of agreement between the IGRAs was observed, they are commonly discordant with the TST. The correct interpretation of a negative assay in a child with a positive skin test in clinical practice remains challenging and highlights the need for longitudinal studies to determine the negative predictive value of IGRAs

Uloga testova otpuštanja interferona gama u nadzoru nad tuberkulozom

Tuberculosis is still one of the major global public health threats. Countries with low incidence must focus on exhausting the reservoir of future cases by preventing reactivation. Therefore, it is important to identify and effectively treat those individuals who have latent tuberculosis infection and who may develop active disease. The tuberculin skin test has been the standard for detection of immune response against M. tuberculosis since the beginning of the 20th century. The new millennium has brought advancement in the diagnosis of latent tuberculosis infection. The name of the new blood test is interferon-gamma release assay (IGRA). Croatia is a middle-incidence country with a long decreasing trend and developed tuberculosis control. To reach low incidence and finally eliminate tuberculosis, its tuberculosis programme needs a more aggressive approach that would include intensive contact investigation and treatment of persons with latent tuberculosis infection. This article discusses the current uses of IGRA and its role in tuberculosis control.Tuberkuloza je i danas jedan od vodećih javnozdravstvenih problema. Zemlje s niskom incidencijom fokusiraju se na iscrpljivanje rezervoara budućih slučajeva sprječavanjem reaktivacije bolesti. To se odnosi na traženje i učinkovito liječenje infi ciranih osoba, primarno onih koje su u riziku od obolijevanja nakon infekcije. Tuberkulinski test je od početka 20. stoljeća bio standard u otkrivanju imunosnog odgovora na kontakt s Mycobacterium tuberculosis. Novo tisućljeće donijelo je određeni napredak u obliku novih testova za dijagnozu latentne tuberkulozne infekcije, krvne testove otpuštanja interferona gama. Hrvatska je zemlja srednje incidencije tuberkuloze s dugogodišnjim silaznim trendom i razvijenim protutuberkuloznim aktivnostima. U težnji prema niskoj incidenciji i u konačnici eliminaciji tuberkuloze potrebne su opsežnije aktivnosti unutar državnog programa nadzora nad tuberkulozom, uključujući intenzivnu obradu kontakata i probir na postojanje latentne tuberkulozne infekcije. Ovaj rad razmatra trenutačnu uporabu IGRE (engl. interferon - gamma release assay) i njezinu ulogu u nadzoru nad tuberkulozom

Whole Blood Interferon-Gamma Responses to Mycobacterium tuberculosis Antigens in Young Household Contacts of Persons with Tuberculosis in Uganda

Due to immunologic immaturity, IFN-gamma-producing T cell responses may be decreased in young children compared to adults, thus we hypothesized that IFN-gamma responses to mycobacterial antigens in household contacts exposed to Mycobacterium tuberculosis (Mtb) would be impaired in young children relative to adults. The objective of this study was to compare whole blood IFN-gamma production in response to mycobacterial antigens between children and adults in Uganda.We studied household contacts of persons with culture-positive pulmonary tuberculosis (TB) enrolled in a cohort study conducted in Kampala, Uganda. Whole blood IFN-gamma production in response to Mtb culture-filtrate antigens was measured by ELISA and compared between infants (<2 years old, n = 80), young children (2 <5 years old, n = 216), older children (5 <15 years old, n = 443) and adults (> or =15 years old, n = 528). We evaluated the relationship between IFN-gamma responses and the tuberculin skin test (TST), and between IFN-gamma responses and epidemiologic factors that reflect exposure to Mtb, and the effect of prior BCG vaccination on IFN-gamma responses. Young household contacts demonstrated robust IFN-gamma responses comparable to those of adults that were associated with TST and known risk factors for infection. There was no effect of prior BCG immunization on the IFN-gamma response.Young children in a TB endemic setting can mount robust IFN-gamma responses generally comparable to those of adults, and as in adults, these responses correlated with the TST and known epidemiologic risk factors for Mtb infection

Genetic Compatibility and Virulence of Reassortants Derived from Contemporary Avian H5N1 and Human H3N2 Influenza A Viruses

The segmented structure of the influenza virus genome plays a pivotal role in its adaptation to new hosts and the emergence of pandemics. Despite concerns about the pandemic threat posed by highly pathogenic avian influenza H5N1 viruses, little is known about the biological properties of H5N1 viruses that may emerge following reassortment with contemporary human influenza viruses. In this study, we used reverse genetics to generate the 63 possible virus reassortants derived from H5N1 and H3N2 viruses, containing the H5N1 surface protein genes, and analyzed their viability, replication efficiency, and mouse virulence. Specific constellations of avian–human viral genes proved deleterious for viral replication in cell culture, possibly due to disruption of molecular interaction networks. In particular, striking phenotypes were noted with heterologous polymerase subunits, as well as NP and M, or NS. However, nearly one-half of the reassortants replicated with high efficiency in vitro, revealing a high degree of compatibility between avian and human virus genes. Thirteen reassortants displayed virulent phenotypes in mice and may pose the greatest threat for mammalian hosts. Interestingly, one of the most pathogenic reassortants contained avian PB1, resembling the 1957 and 1968 pandemic viruses. Our results reveal the broad spectrum of phenotypes associated with H5N1/H3N2 reassortment and a possible role for the avian PB1 in the emergence of pandemic influenza. These observations have important implications for risk assessment of H5N1 reassortant viruses detected in surveillance programs