Irene Najjingo’s research while affiliated with Makerere University and other places

Background: Mechanisms by which prior tuberculosis (TB) increases long-term risk for cancer, cardiovascular, and neurological disorders remain unclear, particularly in people with HIV (PWH). This study investigated DNA methylation (DNAm) patterns and associated pathways in PWH with and without prior TB infection. Methods: DNAm was analyzed in blood samples from 30 PWH (10 with prior latent TB infection [LTBI], 10 with previous successfully treated active TB, and 10 with no TB) using the Illumina MethylationEPIC BeadChip covering over 850,000 CpG sites. Functional enrichment analyses for Gene Ontology, KEGG pathways, and gene set enrichment analysis were performed. Statistical significance was set at a false discovery rate of <0.05. Results: A total of 25,084 differentially methylated CpGs (dmCpGs) were identified in the prior active TB vs. no TB comparison, corresponding to 8 differentially methylated regions (DMRs) in KCNC4-DT, GRAMD1C, ZNF44, FIGN, KCNN3, and PLA2G1B genes. In the LTBI vs. no TB comparison, 7,682 dmCpGs were observed, corresponding to 18 DMRs in SPATC1L, ZFP57, KCNN3, LRSAM1, PLEKHG5, MCF2L, BRSK2, SH3GL2, AP001468.58 and STK32C genes. In both prior active TB vs. no TB and LTBI vs. no TB comparisons, DNAm changes were enriched in pathways related to neurogenesis, neuron differentiation, glutamatergic synapse, and neuroactive ligand-receptor interactions. The LTBI comparison showed additional enrichment in pathways related to synaptic membrane and serotonergic synapse. Cardiovascular pathways were specific to prior active TB, with significant enrichment in vascular smooth muscle contraction, arrhythmogenic right ventricular cardiomyopathy, hypertrophic cardiomyopathy, and dilated cardiomyopathy pathways. Both TB groups showed enrichment in gene sets associated with lung, colorectal, gastric, and breast cancers. The prior active TB group demonstrated additional enrichment for prostate cancer and proteoglycans in cancer, while the LTBI group had additional enrichment for endometrial, esophageal, liver cancers, and Ewing's sarcoma. Conclusion: Prior TB infection in PWH is associated with DNAm changes in pathways related to neural function, cardiovascular health, and cancer risk suggesting epigenetic mechanisms for TB-related long-term complications.

Hematological cancers, including Leukemias and Lymphomas, and their associated chemotherapy and disease-specific factors, are linked to impaired granulocyte function and numbers, increasing the risk of opportunistic infections, often presenting as fever. Human cytomegalovirus (HCMV) is one of the significant opportunistic infections in these patients, but limited data exists on its seroprevalence and active infection burden among febrile hematological cancer patients in Uganda. We conducted a cross-sectional study from June to August 2017 at the Uganda Cancer Institute (UCI). Blood samples from 161 febrile hematological cancer patients were collected. HCMV exposure was assessed using indirect enzyme-linked immunosorbent assay for IgG and IgM antibodies, and active infection was confirmed with PCR testing and gel electrophoresis. IgG positivity indicated previous exposure, while positive IgM or PCR results indicated active infection. Overall, HCMV seroprevalence based on IgG and/or IgM positivity was 106/161 (66%). IgG alone, IgM alone, and combined IgG/IgM positivity prevalence rates were 57/161 (35.4%), 22/161 (13.6%), and 27/161 (16.7%), respectively. HCMV DNA PCR was positive in 5 of the 161 (3%) samples. Among PCR-positive patients, one (20%) was positive for IgG alone, two (40%) for IgM alone, and two (40%) for both IgG and IgM. Active infection based on positive IgM and HCMV DNA PCR was found in 23 of the 161 (14.3%) patients. Two-thirds of febrile patients with hematological malignancies in Uganda had been exposed to HCMV infection, with 14.3% showing active infection. Routine testing for active HCMV infection among febrile hematological cancer patients at the UCI is essential for timely and appropriate antiviral treatment. IMPORTANCE In this paper, we demonstrated that over two-thirds of feverish patients with blood cancers such as leukemia at the Uganda Cancer Institute are already exposed to a type of virus infection called the human cytomegalovirus (HCMV), and 14% of the patients have active disease due to this virus. This was confirmed through finding blood samples testing positive for a type of protective antibody called IgM and also upon virus DNA detection in the blood of those patients. Routine testing for this virus is not usually done in the study settings. Our findings reveal and emphasize the importance of routinely testing blood samples for active infection with this virus among the feverish patients with blood cancers in the study settings, and prompt initiation of antiviral treatment of the actively infected patients

In 2021, an estimated 10.6 million people fell ill with tuberculosis (TB) globally and 11.3% were children. About 40% of children aged five to fourteen years with TB are missed annually. In Uganda, 44% of adolescents with chronic cough of more than two weeks do not seek care from health facilities. Therefore, strategies to promote health care-seeking behaviour among adolescents were urgently needed to resolve the gap. In regard to this, the research project utilized a before and after design, in which the number of adolescents (10-19years) enrolled in the project health facilities were compared before and after the intervention. The intervention package that comprised of tuberculosis awareness and screening information was developed together with adolescents, thus; a human-centred approach was used. The package consisted of TB screening cards, poster messages and a local song. The song was broadcasted in the community radios. Poster messages were deployed in the community by the village health teams (VHTS). The TB screening cards were given to TB positive and presumptive adults to screen adolescents at home. Adolescents that were found with TB symptoms were referred to the project health facilities. Socio-demographic and clinical characteristics of eligible adolescents were collected in a period of six months from Kawolo, Iganga, Gombe and Kiwoko health facilities. To determine the effectiveness of the package, before and after intervention data were equally collected. A total of 394 adolescents were enrolled, majority (76%) were school going. The intervention improved adolescent TB care seeking in the four project health facilities. The average number of adolescents screened increased from 159 to 309 (incidence rate ratio (IRR) = 1.9, P<0.001, 95% CI [1.9, 2.0]). Those presumed to have TB increased from 13 to 29(IRR = 2.2, P<0.001, 95% CI [1.9, 2.5]). The ones tested with GeneXpert increased in average from 8 to 28(IRR = 3.3, P<0.001, 95% CI [2.8, 3.8]). There was a minimal increase in the average monthly number of adolescents with a positive result of 0.8, from 1.6 to 2.4(p = 0.170) and linkage to TB care services of 1.1, from 2 to 3.1(p = 0.154). The project improved uptake of TB services among adolescents along the TB care cascade. We recommend a robust and fully powered randomized controlled trial to evaluate the effectiveness of the Package.

Introduction Dimorphic fungi cause infection following the inhalation of spores into the pulmonary system. In the lower respiratory tract, the conidia transform into yeasts, which are engulfed by alveolar macrophages and may be destroyed without disease manifestation. However, in some immunocompromised individuals, they may persist and cause active fungal disease characterized by formation of granulomas in the infected tissues, which may mimic Mycobacterium tuberculosis (MTB). Objective To determine the prevalence of pulmonary dimorphic fungal infections among HIV/AIDS patients with non‐TB chronic cough at Mulago National Referral and Teaching Hospital in Kampala, Uganda. Methods Sputum samples were collected from 175 consented HIV/AIDS patients attending the immuno‐suppression syndrome (ISS) clinic at the hospital. Upon Xpert MTB/RIF sputum testing, 21 patients tested positive for MTB, and these were excluded from further analysis. The other 154 sputum negative samples were then subjected to PCR for dimorphic fungi at MBN Clinical Laboratories. Singleplex PCR was used to detect the target sequences in selected respective genes of each dimorphic fungal species of interest. DNA amplicons were detected based on gel electrophoresis. Results Dimorphic fungi were detected in 16.2% (25/154) of the studied population. Of these 9.1% (14/154) had Blastomyces dermatitidis and 7.1% (11/154) had Talaromyces marneffei . The remaining 84% of the studied participants had no dimorphic fungi. Histoplasma capsulatum , Coccidioides immitis and Paracoccidioides brasiliensis were not detected in any of the participants. Conclusion Dimorphic fungi ( B. dermatitidis and T. marneffei ) were found in 16.2% of the HIV/AIDS patients with non‐TB chronic cough in Kampala, Uganda. We recommend routine testing for these pathogens among HIV/AIDS patients with chronic cough.

Background Tuberculosis (TB) care could be considered as a continuum from symptom recognition, decision to seek care, diagnosis, treatment initiation and treatment completion, with care along the continuum influenced by several factors. Gender dimensions could influence TB care, and indeed, more men than women are diagnosed with TB each year. The study was done to identify social stratifiers that intersect with gender to influence TB care. Methods A cross-sectional qualitative study was done at four health facilities in 3 districts in central Uganda between October 2020 and December 2020. Data was collected from patients seeking a diagnosis or on TB treatment through focus group discussions and key informant interviews. Key themes around gender guided by a gender and intersectionality lens were developed and thereafter thematic content analysis was done. Results Women have increased vulnerability to TB due to bio mass exposure through roles like cooking. Women have increased access to health care services as they interface with the health care system frequently given their role as child bearers and child care givers. Men have a duty to provide for their families and this most often is prioritised over healthcare seeking, and together with belief that they are powerful beings leads to poor healthcare seeking habits and delays in healthcare seeking. Decisions on when and where to seek care were not straightforward for women, who most often rely on their husbands/partners to make decisions. Conclusions Men and women experience challenges to TB care, and that these challenges are deeply rooted in roles assigned to them and further compounded by masculinity. These challenges need to be addressed through intersectional gender responsive interventions if TB control is to be improved. Graphical Abstract

Asthma is the commonest chronic childhood illness but in young children, it is under/mis-diagnosed leading to avoidable morbidity and mortality. We explored the caregivers’ and health care providers’ understanding and practices around diagnosis and management of asthma in children less than 5 years, and how they contribute to diagnostic delays and inappropriate care of the affected children. Semi-structured interviews with 17 healthcare providers and 8 caregivers were conducted. Data were thematically analysed. Most caregivers described their children as having recurrent cough, difficult breathing, and wheezing, commonly triggered by a common cold and cold weather. They visited several health facilities but were not told the diagnosis and often ineffective antibiotics and cough syrups were prescribed/used. Family social and economic stress and frustration with the healthcare system was common. Healthcare providers never considered asthma in children with recurrent respiratory symptoms but primarily diagnosed pneumonia and prescribed antibiotics. Tuberculos and malaria were suspected in some children with recurrent respiratory illness, and malaria medicine was prescribed. Recurrent respiratory symptoms without respiratory distress were regarded as a simple illness. In conclusion, the study identified major gaps in understanding of asthma symptoms amongst healthcare providers leading to misdiagnosis and mistreatment of children with asthma. Caregivers carefully and comprehensively describe asthma symptoms, but the health system is not responsive to their needs that could help in diagnosis of asthma and appropriate care.

Introduction Dimorphic fungi cause infection following inhalation of spores into the pulmonary system. In the lower respiratory tract, the conidia transform into the yeast phase which are engulfed by alveolar macrophages and may be destroyed without disease manifestation. However, in some cases they may persist and cause fungal disease characterized by formation of granulomas in the infected tissues, which may mimic MTB. Objective To explore if dimorphic fungi play any role in pulmonary disease among XpertTB/RIF Negative HIV Patients with chronic cough attending ISS Clinic at Mulago hospital Uganda. Methods Sputum samples were collected from 175 consented HIV infected patients attending ISS Clinic. Upon Xpert/RIF test at ISS Clinic 21 of these tested positive, the 154 negative sputum samples were then subjected to PCR for dimorphic fungi at MBN Clinical Laboratories. Singleplex PCR using specific primers was used to detect a target sequency in the gene of each dimorphic fungi of interest, the resulting amplicons were electrophoresed on a 2% gel then visualized under UV light. Results Blastomyces dermatitidis and Tarolomyces marneffei were detected in 16.4% of the studied participants, with 9.1% and 7.1% respectively and 83.8% of the participant sample had no dimorphic fungi. Coccidiodes immitis, Paracoccidiodes brasiliensis and Histoplasma capsulatum were not detected in any of the participants Conclusion Dimorphic fungi play a role in pulmonary disease among the HIV/AIDS with non- TB chronic in Uganda.

Introduction In 2021, an estimated 10.6 million people fell ill with tuberculosis (TB) globally, 1.2 million of these were children. About 40% of them aged between 5 and 14 years with TB are missed annually. In Uganda, 44% of adolescents with chronic cough of >=2 weeks do not seek care from health facilities. Therefore, strategies to promote health care-seeking behaviour among adolescents are urgently needed. We piloted a project (TEEN TB project) aimed at improving uptake of tuberculosis (TB) care services among adolescents at Ugandan health facilities. Methodology We developed an adolescent TB awareness and screening package using the human centred design. This technique puts real people at the centre of the development process. The package consisted of 3 interventions (TB screening cards, adolescent-TB awareness poster messages and a local TB awareness song) deployed in the project health facilities and their surrounding communities. Data on socio-demographic and clinical characteristics of adolescents were collected for the period between October 2021 and March 2022 at 4 project health facilities (Kawolo, Iganga, Gombe and Kiwoko). We collected before and after intervention data from facility level records to determine the effect of the package. Results A total of 394 adolescents were included and the majority (76%) were still in school. Overall, the intervention improved adolescent TB care in the four project health facilities. The average number of adolescents screened increased by 94% from 159 to 309, with an incidence rate ratio (IRR) of 1.9 ( 95% CI: 1.9- 2.0, p <0.001), there was a 2-fold increase among those presumed to have TB; from 13 to 29, IRR of 2.2 (95% CI: 1.9-2.5, p <0.001) and those tested with GeneX-pert and microscopy increased more than 3 times from 8 to 28, IRR of 3.3 ( 95% CI: 2.8-3.8, p <0.001). There was a minimal increase in the average monthly number of adolescents with a positive result from 1.6 to 2.4 and linkage to TB care services from 2 to 3.1. These were not statistically significant at p=0.170 and p=0.154 respectively. Conclusion The project improved uptake of TB services among adolescents along the TB care cascade (screening, TB testing and linkage to care). We recommend a robust and fully powered randomized controlled trial to evaluate the effectiveness of the package.

Introduction Tuberculosis (TB) remains a major cause of morbidity and mortality, especially in sub-Saharan Africa. We qualitatively evaluated the implementation of an Evidence-Based Multiple Focus Integrated Intensified TB Screening package (EXIT-TB) in the East African region, aimed at increasing TB case detection and number of patients receiving care. Objective We present the accounts of participants from Tanzania, Kenya, Uganda, and Ethiopia regarding the implementation of EXIT-TB, and suggestions for scaling up. Methods A qualitative descriptive design was used to gather insights from purposefully selected healthcare workers, community health workers, and other stakeholders. A total of 27, 13, 14, and 19 in-depth interviews were conducted in Tanzania, Kenya, Uganda, and Ethiopia respectively. Data were transcribed and translated simultaneously and then thematically analysed. Results The EXIT-TB project was described to contribute to increased TB case detection, improved detection of Multidrug-resistant TB patients, reduced delays and waiting time for diagnosis, raised the index of TB suspicion, and improved decision-making among HCWs. The attributes of TB case detection were: (i) free X-ray screening services; (ii) integrating TB case-finding activities in other clinics such as Reproductive and Child Health clinics (RCH), and diabetic clinics; (iii), engagement of CHWs, policymakers, and ministry level program managers; (iv) enhanced community awareness and linkage of clients; (v) cooperation between HCWs and CHWs, (vi) improved screening infrastructure, (vii) the adoption of the new simplified screening criteria and (viii) training of implementers. The supply-side challenges encountered ranged from disorganized care, limited space, the COVID-19 pandemic, inadequate human resources, inadequate knowledge and expertise, stock out of supplies, delayed maintenance of equipment, to absence of X-ray and GeneXpert machines in some facilities. The demand side challenges ranged from delayed care seeking, inadequate awareness, negative beliefs, fears towards screening, to financial challenges. Suggestions for scaling up ranged from improving service delivery, access to diagnostic equipment and supplies, and infrastructure, to addressing client fears and stigma. Conclusion The EXIT-TB package appears to have contributed towards increasing TB case detection and reducing delays in TB treatment in the study settings. Addressing the challenges identified is needed to maximize the impact of the EXIT-TB intervention.