Impaired resolution of blood transcriptomes through tuberculosis treatment with diabetes comorbidity.

BACKGROUND: People with diabetes are more likely to develop tuberculosis (TB) and to have poor TB-treatment outcomes than those without. We previously showed that blood transcriptomes in people with TB-diabetes (TB-DM) co-morbidity have excessive inflammatory and reduced interferon responses at diagnosis. It is unknown whether this persists through treatment and contributes to the adverse outcomes. METHODS: Pulmonary TB patients recruited in South Africa, Indonesia and Romania were classified as having TB-DM, TB with prediabetes, TB-related hyperglycaemia or TB-only, based on glycated haemoglobin concentration at TB diagnosis and after 6 months of TB treatment. Gene expression in blood at diagnosis and intervals throughout treatment was measured by unbiased RNA-Seq and targeted Multiplex Ligation-dependent Probe Amplification. Transcriptomic data were analysed by longitudinal mixed-model regression to identify whether genes were differentially expressed between clinical groups through time. Predictive models of TB-treatment response across groups were developed and cross-tested. RESULTS: Gene expression differed between TB and TB-DM patients at diagnosis and was modulated by TB treatment in all clinical groups but to different extents, such that differences remained in TB-DM relative to TB-only throughout. Expression of some genes increased through TB treatment, whereas others decreased: some were persistently more highly expressed in TB-DM and others in TB-only patients. Genes involved in innate immune responses, anti-microbial immunity and inflammation were significantly upregulated in people with TB-DM throughout treatment. The overall pattern of change was similar across clinical groups irrespective of diabetes status, permitting models predictive of TB treatment to be developed. CONCLUSIONS: Exacerbated transcriptome changes in TB-DM take longer to resolve during TB treatment, meaning they remain different from those in uncomplicated TB after treatment completion. This may indicate a prolonged inflammatory response in TB-DM, requiring prolonged treatment or host-directed therapy for complete cure. Development of transcriptome-based biomarker signatures of TB-treatment response should include people with diabetes for use across populations.

Transcriptional profiles predict treatment outcome in patients with tuberculosis and diabetes at diagnosis and at two weeks after initiation of anti-tuberculosis treatment.

BACKGROUND: Globally, the tuberculosis (TB) treatment success rate is approximately 85%, with treatment failure, relapse and death occurring in a significant proportion of pulmonary TB patients. Treatment success is lower among people with diabetes mellitus (DM). Predicting treatment outcome early after diagnosis, especially in TB-DM patients, would allow early treatment adaptation for individuals and may improve global TB control. METHODS: Samples were collected in a longitudinal cohort study of adult TB patients from South Africa (n  =  94) and Indonesia (n  =  81), who had concomitant DM (n  =  59), intermediate hyperglycaemia (n  =  79) or normal glycaemia/no DM (n  =  37). Treatment outcome was monitored, and patients were categorized as having a good (cured) or poor (failed, recurrence, died) outcome during treatment and 12 months follow-up. Whole blood transcriptional profiles before, during and at the end of TB treatment were characterized using unbiased RNA-Seq and targeted gene dcRT-MLPA. FINDINGS: We report differences in whole blood transcriptome profiles, which were observed before initiation of treatment and throughout treatment, between patients with a good versus poor TB treatment outcome. An eight-gene and a 22-gene blood transcriptional signature distinguished patients with a good TB treatment outcome from patients with a poor TB treatment outcome at diagnosis (AUC = 0·815) or two weeks (AUC = 0·834) after initiation of TB treatment, respectively. High accuracy was obtained by cross-validating this signature in an external cohort (AUC = 0·749). INTERPRETATION: These findings suggest that transcriptional profiles can be used as a prognostic biomarker for treatment failure and success, even in patients with concomitant DM. FUNDING: The research leading to these results, as part of the TANDEM Consortium, received funding from the European Community's Seventh Framework Programme (FP7/2007-2013 Grant Agreement No. 305279) and the Netherlands Organization for Scientific Research (NWO-TOP Grant Agreement No. 91214038). The research leading to the results presented in the Indian validation cohort was supported by Research Council of Norway Global Health and Vaccination Research (GLOBVAC) projects: RCN 179342, 192534, and 248042, the University of Bergen (Norway).

Vaginal microbiota in ethnically diverse young women who did or did not develop pelvic inflammatory disease: community-based prospective study.

OBJECTIVES: A lactobacilli-dominated vaginal microbiome may protect against pelvic inflammatory disease (PID), but one dominated by Gardnerella species might increase susceptibility. Not all lactobacilli are equally protective. Recent research suggests that D(-) isomer lactic acid producing lactobacilli (Lactobacillus crispatus, Lactobacillus jensenii and Lactobacillus gasseri) may protect against infection with Chlamydia trachomatis, an important cause of PID. Lactobacillus iners , which produces L(+) isomer lactic acid, may be less protective. We investigated the microbiome in stored vaginal samples from participants who did or did not develop PID during the prevention of pelvic infection (POPI) chlamydia screening trial. METHODS: Long-read 16S rRNA gene nanopore sequencing was used on baseline vaginal samples (one per participant) from all 37 women who subsequently developed clinically diagnosed PID during 12-month follow-up, and 111 frequency matched controls who did not, matched on four possible risk factors for PID: age <20 versus ≥20, black ethnicity versus other ethnicity, chlamydia positive versus negative at baseline and ≥2 sexual partners in the previous year versus 0-1 partners. RESULTS: Samples from 106 women (median age 19 years, 40% black ethnicity, 22% chlamydia positive, 54% reporting multiple partners) were suitable for analysis. Three main taxonomic clusters were identified dominated by L. iners, L. crispatus and Gardnerella vaginalis. There was no association between a more diverse, G. vaginalis dominated microbiome and subsequent PID, although increased Shannon diversity was associated with black ethnicity (p=0.002) and bacterial vaginosis (diagnosed by Gram stain p<0.0001). Women who developed PID had similar relative abundance of protective D(-) isomer lactic acid producing lactobacilli to women without PID, but numbers of PID cases were small. CONCLUSIONS: In the first-ever community-based prospective study of PID, there was no clear association between the vaginal microbiome and subsequent development of PID. Future studies using serial samples may identify vaginal microbial communities that may predispose to PID.

Alcohol use, cigarette smoking, vaping and number of sexual partners: A cross-sectional study of sexually active, ethnically diverse, inner city adolescents.

CONTEXT: There are few UK data on the prevalence and clustering of risky behaviours in ethnically diverse adolescents. OBJECTIVES: To investigate the prevalence of reported alcohol use, smoking and vaping, and explore whether these behaviours are associated with increased numbers of sexual partners. DESIGN: Questionnaire survey of 'Test n Treat' chlamydia screening trial participants. SETTING AND PARTICIPANTS: Sexually active students attending six London technical colleges completed confidential questionnaires and provided genitourinary samples. RESULTS: The median age of the 509 participants was 17 years (IQR: 16-18), 47% were male, 50% were of black ethnicity, 55% reported ≥2 sexual partners in the past year (67% of males and 45% of females) and 6.2% had chlamydia infection and 0.6% gonorrhoea. Almost half (48%) reported getting drunk in the past month, 33% smoked cigarettes and 7% had ever vaped. A larger percentage of students with ≥2 sexual partners than 0-1 partners reported getting drunk in the past month (53.7%, 144/268% versus 42.2% 94/223, adjusted prevalence ratio: 1.33, 95% confidence interval: 1.11-1.61) and smoking cigarettes (36.6%, 100/273% versus 30.2%, 67/222, 1.34 (1.05-1.70)). By contrast, multiple sexual partners were not associated with vaping or chlamydia infection, but numbers were small. CONCLUSIONS: We found high prevalences of risky behaviour and an association between multiple sexual partners and smoking and/or getting drunk. Findings support the introduction of compulsory sex and relationship education in UK secondary schools, including information about the adverse effects of alcohol and smoking. PUBLIC CONTRIBUTION: Participants helped with study design, conduct and interpretation.

Impact of Intermediate Hyperglycemia and Diabetes on Immune Dysfunction in Tuberculosis.

BACKGROUND: People with diabetes have an increased risk of developing active tuberculosis (TB) and are more likely to have poor TB-treatment outcomes, which may impact on control of TB as the prevalence of diabetes is increasing worldwide. Blood transcriptomes are altered in patients with active TB relative to healthy individuals. The effects of diabetes and intermediate hyperglycemia (IH) on this transcriptomic signature were investigated to enhance understanding of immunological susceptibility in diabetes-TB comorbidity. METHODS: Whole blood samples were collected from active TB patients with diabetes (glycated hemoglobin [HbA1c] ≥6.5%) or IH (HbA1c = 5.7% to <6.5%), TB-only patients, and healthy controls in 4 countries: South Africa, Romania, Indonesia, and Peru. Differential blood gene expression was determined by RNA-seq (n = 249). RESULTS: Diabetes increased the magnitude of gene expression change in the host transcriptome in TB, notably showing an increase in genes associated with innate inflammatory and decrease in adaptive immune responses. Strikingly, patients with IH and TB exhibited blood transcriptomes much more similar to patients with diabetes-TB than to patients with only TB. Both diabetes-TB and IH-TB patients had a decreased type I interferon response relative to TB-only patients. CONCLUSIONS: Comorbidity in individuals with both TB and diabetes is associated with altered transcriptomes, with an expected enhanced inflammation in the presence of both conditions, but also reduced type I interferon responses in comorbid patients, suggesting an unexpected uncoupling of the TB transcriptome phenotype. These immunological dysfunctions are also present in individuals with IH, showing that altered immunity to TB may also be present in this group. The TB disease outcomes in individuals with IH diagnosed with TB should be investigated further.

Human papillomavirus (HPV) vaccination and oropharyngeal HPV in ethnically diverse, sexually active adolescents: community-based cross-sectional study.

OBJECTIVES: Oropharyngeal squamous cell carcinoma is the most common human papillomavirus (HPV)-associated cancer in the UK, but little is known about the prevalence of oropharyngeal HPV in sexually active teenagers. We investigated reported HPV vaccination coverage (in females) and prevalence of oropharyngeal HPV in sexually active students attending six technical colleges in London, UK. METHODS: In 2017, we obtained mouthwash samples and questionnaires from male and female students taking part in the 'Test n Treat' chlamydia screening trial. Samples were subjected to HPV genotyping. RESULTS: Of 232 participants approached, 202 (87%) provided a mouthwash sample and questionnaire. Participants' median age was 17 years and 47% were male. Most (73%) were from black and minority ethnic groups, 64% gave a history of oral sex, 52% reported having a new sexual partner in the past 6 months, 33% smoked cigarettes, 5.9% had concurrent genitourinary Chlamydia trachomatis infection and 1.5% Neisseria gonorrhoeae and 5.0% were gay or bisexual. Only 47% (50/107) of females reported being vaccinated against HPV 16/18, of whom 74% had received ≥2 injections. HPV genotyping showed three mouthwash samples (1.5%, 95% CI 0.3% to 4.3%) were positive for possible high-risk human papillomavirus (HR-HPV), one (0.5%, 0.0% to 2.7%) for low-risk HPV 6/11, but none (0.0%, 0.0% to 1.8%) for HR-HPV. Four samples (2.0%, 0.5% to 5.0%) were positive for HPV16 using a HPV16 type-specific quantitative PCR, but these were at a very low copy number and considered essentially negative. CONCLUSIONS: Despite the high prevalence of oral sex and genitourinary chlamydia and low prevalence of HPV vaccination, the prevalence of oropharyngeal HR-HPV in these adolescents was negligible.

Understanding the acceptability, barriers and facilitators for chlamydia and gonorrhoea screening in technical colleges: qualitative process evaluation of the "Test n Treat" trial.

BACKGROUND: Low uptake of sexually transmitted infection testing by sexually active young people is a worldwide public health problem. Screening in non-medical settings has been suggested as a method to improve uptake. The "Test n Treat" feasibility trial offered free, on-site rapid chlamydia/gonorrhoea tests with same day treatment for chlamydia (and gonorrhoea treatment at a local clinic,) to sexually active students (median age 17 years) at six technical colleges in London. Despite high rates of chlamydia (6% prevalence), uptake of testing was low (< 15%). In a qualitative study we explored the acceptability, including barriers and facilitators to uptake, of on-site chlamydia screening. METHODS: In 2016-17 we conducted a qualitative study in the interpretative tradition using face to face or telephone semi-structured interviews with students (n = 26), teaching staff (n = 3) and field researchers (n = 4). Interviews were digitally recorded, transcribed and thematically analysed. RESULTS: From the student perspective, feelings of embarrassment and the potential for stigma were deterrents to sexually transmitted infection testing. While the non-medical setting was viewed as mitigating against stigma, for some students volunteering to be screened exposed them to detrimental judgements by their peers. A small financial incentive to be screened was regarded as legitimising volunteering in a non-discrediting way. Staff and researchers confirmed these views. The very low level of knowledge about sexually transmitted infections influenced students to not view themselves as candidates for testing. There were also suggestions that some teenagers considered themselves invulnerable to sexually transmitted infections despite engaging in risky sexual behaviours. Students and researchers reported the strong influence peers had on uptake, or not, of sexually transmitted infection testing. CONCLUSIONS: This study offers new insights into the acceptability of college-based sexually transmitted infection screening to young, multi-ethnic students. Future studies in similar high risk, hard to reach groups should consider linking testing with education about sexually transmitted infections, offering non stigmatising incentives and engaging peer influencers.

Near patient chlamydia and gonorrhoea screening and treatment in further education/technical colleges: a cost analysis of the 'Test n Treat' feasibility trial.

BACKGROUND: Community-based screening may be one solution to increase testing and treatment of sexually transmitted infections in sexually active teenagers, but there are few data on the practicalities and cost of running such a service. We estimate the cost of running a 'Test n Treat' service providing rapid chlamydia (CT) and gonorrhoea (NG) testing and same day on-site CT treatment in technical colleges. METHODS: Process data from a 2016/17 cluster randomised feasibility trial were used to estimate total costs and service uptake. Pathway mapping was used to model different uptake scenarios. Participants, from six London colleges, provided self-taken genitourinary samples in the nearest toilet. Included in the study were 509 sexually active students (mean 85/college): median age 17.9 years, 49% male, 50% black ethnicity, with a baseline CT and NG prevalence of 6 and 0.5%, respectively. All participants received information about CT and NG infections at recruitment. When the Test n Treat team visited, participants were texted/emailed invitations to attend for confidential testing. Three colleges were randomly allocated the intervention, to host (non-incentivised) Test n Treat one and four months after baseline. All six colleges hosted follow-up Test n Treat seven months after baseline when students received a £10 incentive (to participate). RESULTS: The mean non-incentivised daily uptake per college was 5 students (range 1 to 17), which cost £237 (range £1082 to £88) per student screened, and £4657 (range £21,281 to £1723) per CT infection detected, or £13,970 (range £63,842 to £5169) per NG infection detected. The mean incentivised daily uptake was 19 students which cost £91 per student screened, and £1408/CT infection or £7042/NG infection detected. If daily capacity for screening were achieved (49 students/day), costs including incentives would be £47 per person screened and £925/CT infection or £2774/NG infection detected. CONCLUSIONS: Delivering non-incentivised Test n Treat in technical colleges is more expensive per person screened than CT and NG screening in clinics. Targeting areas with high infection rates, combined with high, incentivised uptake could make costs comparable. TRIAL REGISTRATION: ISRCTN58038795, Assigned August 2016, registered prospectively.

Accuracy of diabetes screening methods used for people with tuberculosis, Indonesia, Peru, Romania, South Africa.

OBJECTIVE: To evaluate the performance of diagnostic tools for diabetes mellitus, including laboratory methods and clinical risk scores, in newly-diagnosed pulmonary tuberculosis patients from four middle-income countries. METHODS: In a multicentre, prospective study, we recruited 2185 patients with pulmonary tuberculosis from sites in Indonesia, Peru, Romania and South Africa from January 2014 to September 2016. Using laboratory-measured glycated haemoglobin (HbA1c) as the gold standard, we measured the diagnostic accuracy of random plasma glucose, point-of-care HbA1c, fasting blood glucose, urine dipstick, published and newly derived diabetes mellitus risk scores and anthropometric measurements. We also analysed combinations of tests, including a two-step test using point-of-care HbA1cwhen initial random plasma glucose was ≥ 6.1 mmol/L. FINDINGS: The overall crude prevalence of diabetes mellitus among newly diagnosed tuberculosis patients was 283/2185 (13.0%; 95% confidence interval, CI: 11.6-14.4). The marker with the best diagnostic accuracy was point-of-care HbA1c (area under receiver operating characteristic curve: 0.81; 95% CI: 0.75-0.86). A risk score derived using age, point-of-care HbA1c and random plasma glucose had the best overall diagnostic accuracy (area under curve: 0.85; 95% CI: 0.81-0.90). There was substantial heterogeneity between sites for all markers, but the two-step combination test performed well in Indonesia and Peru. CONCLUSION: Random plasma glucose followed by point-of-care HbA1c testing can accurately diagnose diabetes in tuberculosis patients, particularly those with substantial hyperglycaemia, while reducing the need for more expensive point-of-care HbA1c testing. Risk scores with or without biochemical data may be useful but require validation.

Disease characteristics and treatment of patients with diabetes mellitus attending government health services in Indonesia, Peru, Romania and South Africa.

OBJECTIVE: To describe the characteristics and management of Diabetes mellitus (DM) patients from low- and middle-income countries (LMIC). METHODS: We systematically characterised consecutive DM patients attending public health services in urban settings in Indonesia, Peru, Romania and South Africa, collecting data on DM treatment history, complications, drug treatment, obesity, HbA1c and cardiovascular risk profile; and assessing treatment gaps against relevant national guidelines. RESULTS: Patients (median 59 years, 62.9% female) mostly had type 2 diabetes (96%), half for >5 years (48.6%). Obesity (45.5%) and central obesity (females 84.8%; males 62.7%) were common. The median HbA1c was 8.7% (72 mmol/mol), ranging from 7.7% (61 mmol/mol; Peru) to 10.4% (90 mmol/mol; South Africa). Antidiabetes treatment included metformin (62.6%), insulin (37.8%), and other oral glucose-lowering drugs (34.8%). Disease complications included eyesight problems (50.4%), EGFR <60 ml/min (18.9%), heart disease (16.5%) and proteinuria (14.7%). Many had an elevated cardiovascular risk with elevated blood pressure (36%), LDL (71.0%) and smoking (13%), but few were taking antihypertensive drugs (47.1%), statins (28.5%) and aspirin (30.0%) when indicated. Few patients on insulin (8.0%), statins (8.4%) and antihypertensives (39.5%) reached treatment targets according to national guidelines. There were large differences between countries in terms of disease profile and medication use. CONCLUSION: DM patients in government clinics in four LMIC with considerable growth of DM have insufficient glycaemic control, frequent macrovascular and other complications, and insufficient preventive measures for cardiovascular disease. These findings underline the need to identify treatment barriers and secure optimal DM care in such settings.

'Test n Treat (TnT)': a cluster-randomised feasibility trial of frequent, rapid-testing and same-day, on-site treatment to reduce rates of chlamydia in high-risk further education college students: statistical analysis plan.

BACKGROUND: There are high rates of sexually transmitted infections (STIs) in ethnically diverse, sexually active students aged 16-24 years attending London further education (FE) colleges. However, uptake of chlamydia screening remains low. The TnT study aims to assess the feasibility of conducting a future trial in FE colleges to investigate if frequent, rapid, on-site testing and treatment (TnT) reduces chlamydia rates. This article presents the statistical analysis plan for the main study publication as approved and signed off by the Trial Management Group prior to the first data extraction for the final report. METHODS/DESIGN: TnT is a cluster-randomised feasibility trial conducted over 7 months with parallel qualitative and economic assessments. Colleges will be randomly allocated into the intervention (TnT) or the control group (no TnT). Six FE colleges in London will be included. At each college for 2 days, 80 consecutive sexually active students aged 16-24 years (total 480 students across all six colleges) will be recruited from public areas and asked to provide baseline samples. One and 4 months after recruitment intervention colleges will be visited on two consecutive days by the TnT team where participating students will be texted and invited to come for same-day, on-site, rapid chlamydia testing and, if positive, treatment. Participants in the control colleges will receive 'thank you' texts 1 and 4 months after recruitment. Seven months after recruitment, participants from both groups will be invited to complete questionnaires and provide samples for TnT. All samples will be tested, and same-day treatment offered to participants with positive results. Key feasibility outcomes include: recruitment rates, testing and treatment uptake rates (at 1 and 4 months) and follow-up rates (at 7 months). TRIAL REGISTRATION: ISRCTN 58038795 . Registered on 31 August 2016.

'Test n Treat (TnT)'- Rapid testing and same-day, on-site treatment to reduce rates of chlamydia in sexually active further education college students: study protocol for a cluster randomised feasibility trial.

BACKGROUND: Sexually active young people attending London further education (FE) colleges have high rates of chlamydia, but screening rates are low. We will conduct a cluster randomised feasibility trial of frequent, rapid, on-site chlamydia testing and same-day treatment (Test and Treat (TnT)) in six FE colleges (with parallel qualitative and economic assessments) to assess the feasibility of conducting a future trial to investigate if TnT reduces chlamydia rates. METHODS: We will recruit 80 sexually active students aged 16-24 years from public areas at each of six colleges. All participants (total n = 480) will be asked to provide samples (urine for males, self-taken vaginal swabs for females) and complete questionnaires on sexual lifestyle and healthcare use at baseline and after 7 months. Participants will be informed that baseline samples will not be tested for 7 months and be advised to get screened separately. Colleges will be randomly allocated to the intervention (TnT) or the control group (no TnT). One and 4 months after recruitment, participants at each intervention college (n = 3) will be texted and invited for on-site chlamydia tests using the 90-min Cepheid GeneXpert system. Students with positive results will be asked to see a visiting nurse health adviser for same-day treatment and partner notification, (backed by genitourinary medicine follow-up). Participants in control colleges (n = 3) will receive 'thank you' texts 1 and 4 months after recruitment. Seven months after recruitment, participants from both groups will be invited to complete questionnaires and provide samples for TnT. All samples will be tested, and same-day treatment offered to students with positive results. Acceptability of TnT will be assessed by qualitative interviews of purposively sampled students (n = 30) and college staff (n = 12). We will collect data on costs of TnT and usual healthcare. DISCUSSION: Findings will provide key values to inform feasibility, sample size and timescales of a future definitive trial of TnT in FE colleges, including: Recruitment rates TnT uptake rates Follow-up rates Prevalence of chlamydia in participants at baseline and 7 months Acceptability of TnT to students and college staff Estimate of the cost per person screened/treated in TnT versus usual care TRIAL REGISTRATION: International Standard Randomised Controlled Trials Registry, ID: ISRCTN58038795 , Registered on 31 August 2016.

The contribution of physical fitness to individual and ethnic differences in risk markers for type 2 diabetes in children: The Child Heart and Health Study in England (CHASE).

BACKGROUND: The relationship between physical fitness and risk markers for type 2 diabetes (T2D) in children and the contribution to ethnic differences in these risk markers have been little studied. We examined associations between physical fitness and early risk markers for T2D and cardiovascular disease in 9- to 10-year-old UK children. METHODS: Cross-sectional study of 1445 9- to 10-year-old UK children of South Asian, black African-Caribbean and white European origin. A fasting blood sample was used for measurement of insulin, glucose (from which homeostasis model assessment [HOMA]-insulin resistance [IR] was derived), glycated hemoglobin (HbA1c), urate, C-reactive protein (CRP), and lipids. Measurements of blood pressure (BP) and fat mass index (FMI) were made; physical activity was measured by accelerometry. Estimated VO2 max was derived from a submaximal fitness step test. Associations were estimated using multilevel linear regression. RESULTS: Higher VO2 max was associated with lower FMI, insulin, HOMA-IR, HbA1c, glucose, urate, CRP, triglycerides, LDL-cholesterol, BP and higher HDL-cholesterol. Associations were reduced by adjustment for FMI, but those for insulin, HOMA-IR, glucose, urate, CRP, triglycerides and BP remained statistically significant. Higher levels of insulin and HOMA-IR in South Asian children were partially explained by lower levels of VO2max compared to white Europeans, accounting for 11% of the difference. CONCLUSIONS: Physical fitness is associated with risk markers for T2D and CVD in children, which persist after adjustment for adiposity. Higher levels of IR in South Asians are partially explained by lower physical fitness levels compared to white Europeans. Improving physical fitness may provide scope for reducing risks of T2D.