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Background Of the millions of people infected with Mycobacterium tuberculosis (Mtb) each year, only a small proportion will develop TB disease, with most immunologically containing or clearing Mtb. Factors influencing TB progression risk are incompletely understood. Co-infections, including influenza, have been proposed as a risk factor for TB progression via disruption of anti-Mtb immune responses. We employed a human influenza challenge study to investigate the effect of systemic influenza infection on host mycobacterial control. Methods A whole blood (WB) mycobacterial growth inhibition assay was utilised to compare mycobacterial growth and anti-mycobacterial immune responses before and after influenza infection. Thirty adults were per-nasally inoculated with H3N2 influenza virus (Day [D] 0). Influenza PCR assay of D4 nasal swab confirmed infection. WB, collected pre- (D0, “pre-influenza”) and post-inoculation (D6, “post-influenza”), was infected with Mycobacterium bovis Bacille Calmette Guerin (BCG)-lux, incubated for 72 hours (h) and WB mycobacterial growth (growth ratio [GR]) measured. In parallel, BCG-lux-infected and uninfected blood aliquots were incubated for 0, 6, 24 and 72 h and measurements of cytokines (Meso Scale) and gene expression (RNA-Sequencing) undertaken. Comparisons between pre- and post-influenza infection blood samples were made (Fig. 1). Blood was aliquoted into triplicate for each timepoint (0 hours [h], 72 h), diluted 1:1 with Gibco Roswell Park Memorial Institute 1640 Medium (RPMI), inoculated with a fixed quantity of BCG-lux and incubated at 37°C in a rocker-incubator. Previous experiments had confirmed and quantified the correlation between BCG-lux luminescence (measured in Relative Light Units [RLU] by a luminometer) and number of mycobacterial colony forming units. At 0 h and 72 h, the luminescence of each triplicate sample was measured in duplicate. The growth ratio (GR) was calculated by division of the median 72 h luminescence value by the median 0 h luminescence value. In parallel, BCG-lux inoculum or sterile PBS were added to additional whole blood aliquots (to give infected and uninfected aliquots), which were incubated at 37°C in a rocker-incubator. At timepoints 0 h, 6 h, 24 h, and 72 h, aliquots were removed from the incubator and centrifuged. Supernatants were frozen and stored for future cytokine analyses (quantification using Meso Scale Discovery [MSD] U-plex). Cell pellets were stabilised and stored for future RNA-sequencing and transcriptomic analysis. Figure created with BioRender.com. Results In 22 influenza PCR-positive (+) subjects, median GR was significantly higher in post- (1.69) vs pre-influenza samples (1.03, p=0.0016) with no significant difference in the PCR-negative subjects (Fig. 2). Significant differences in BCG-lux-stimulated cytokine production were observed between post- and pre-influenza samples in PCR+ subjects (Fig. 3). Transcriptomic analysis identified significantly differentially expressed genes in the post- vs pre-influenza samples and differences between the groups over time, mapping to pathways related to TB susceptibility. Complete BCG-lux growth ratio (GR) data were available for 28/30 participants. Comparisons of median GRs pre- and post-influenza infection for (A) 22 participants who were influenza PCR-positive on day 4 and (B) 6 participants who were influenza PCR-negative on day 4. Significance (paired analysis): ** p≤0.01; ns p>0.05. (C) Change in GR from pre-influenza baseline was calculated for each individual (GR-Post divided by GR-Pre, expressed as a percentage) and compared between the influenza PCR-positive and PCR-negative participants. Significance: ** p≤0.01. Cytokine concentrations were measured in duplicate in supernatants from blood incubated with BCG-lux for 0 hours (h), 6 h, 24 h and 72 h, using Meso Scale Discovery U-plex. Results are available for a subset of 15 participants who had influenza infection confirmed by influenza PCR-positive assay on day 4 nasal swab. Comparisons of median concentrations between pre- and post-influenza samples and individual participants’ paired samples are shown for (A, B) Interleukin-1β (IL-1β) at 24 h; (C, D) Tumour necrosis factor-α (TNF-α) at 6 h; (E, F) Interleukin-10 (IL-10) at 0 h; (G, H) IL-10 at 24 h. Significance (paired analysis): **** p≤0.0001; ** p≤0.01. Conclusion Systemic influenza infection reduces whole blood control of mycobacteria through modulation of anti-mycobacterial immune responses. Cytokine differences may contribute. These novel findings suggest that influenza may be a risk factor for TB disease and influenza vaccine could have a non-specific effect on TB immunity. Disclosures Christopher W. Woods, MD MPH, Predigen, Inc: Co-founder Christopher Chiu, BMBCh PhD, GlaxoSmithKline: Grant/Research Support|Merck: Grant/Research Support|Reithera: Advisor/Consultant.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is associated with highly variable clinical outcomes. Studying the temporal dynamics of host whole blood gene expression during SARS-CoV-2 infection can elucidate the biological processes that underlie these diverse clinical phenotypes. We employed a novel pseudotemporal approach using MaSigPro to model and compare the trajectories of whole blood transcriptomic responses in patients with mild, moderate and severe COVID-19 disease. We identified 5,267 genes significantly differentially expressed (SDE) over pseudotime and between severity groups and clustered these genes together based on pseudotemporal trends. Pathway analysis of these gene clusters revealed upregulation of multiple immune, coagulation, platelet and senescence pathways with increasing disease severity and downregulation of T cell, transcriptional and cellular metabolic pathways. The gene clusters exhibited differing pseudotemporal trends. Monoamine oxidase B was the top SDE gene, upregulated in severe>moderate>mild COVID-19 disease. This work provides new insights into the diversity of the host response to SARS-CoV-2 and disease severity and highlights the utility of pseudotemporal approaches in studying evolving immune responses to infectious diseases.

We investigated the longitudinal whole blood transcriptional profile responses to tuberculosis preventive therapy of 18 IGRA-positive tuberculosis contacts and IGRA-negative, tuberculosis-unexposed healthy controls.Longitudinal unsupervised clustering analysis with a subset of 474 most variable genes in antigen-stimulated blood separated the IGRA+ participants into two distinct subgroups, one of which clustered with the IGRA-negative controls. 117 probes were significantly differentially expressed over time between the two cluster groups, many of them associated with immunological pathways important in mycobacterial control.We contend that the differential host RNA response reflects lack of M.tuberculosis (Mtb) viability in the group that clustered with the IGRA-unexposed healthy controls, and Mtb viability in the group (1/3 of IGRA-positives) that clustered away.Gene expression patterns in the blood of IGRA+ individuals emerging during the course of PT, which reflect Mtb viability, could have major implications in the identification of risk of progression, treatment stratification and biomarker development.

Supplementary data: available online at https://www.sciencedirect.com/science/article/pii/S1472979220302006?via%3Dihub#appsec1 Copyright © 2020 The Authors. We hypothesised that individuals with immunological sensitisation to Mycobacterium tuberculosis (Mtb), conventionally regarded as evidence of latent tuberculosis infection (LTBI), would demonstrate binary responses to preventive therapy (PT), reflecting the differential immunological consequences of the sterilisation of viable infection in those with active Mtb infection versus no Mtb killing in those who did not harbour viable bacilli. We investigated longitudinal whole blood transcriptional profile responses to PT of Interferon gamma release assay (IGRA)-positive tuberculosis contacts and IGRA-negative, tuberculosis-unexposed controls. Longitudinal unsupervised clustering analysis with a subset of 474 most variable genes in antigen-stimulated blood separated the IGRA-positive participants into two distinct subgroups, one of which clustered with the IGRA-negative controls. 117 probes were differentially expressed over time between the two cluster groups, many of them associated with immunological pathways important in mycobacterial control. We contend that the differential host RNA response reflects lack of Mtb viability in the group that clustered with the IGRA-negative unexposed controls, and Mtb viability in the group (1/3 of IGRA-positives) that clustered away. Gene expression patterns in the blood of IGRA-positive individuals emerging during the course of PT, which reflect Mtb viability, could have major implications in the identification of risk of progression, treatment stratification and biomarker development. This work was supported by a British Infection Association Small Project Research Grant (2016) and a Rosetrees Trust Seed Corn Award (# JS15/M660). C.B. was funded by an Academic Clinical Fellowship from the National Institute for Health Research (NIHR) (ACF-2012-18-008) and currently receives support from an Imperial 4i Wellcome Trust/NIHR Imperial BRC Clinical PhD Fellowship. M.K. receives support from the NIHR Imperial College BRC and the Wellcome Trust (Sir Henry Wellcome Fellowship grant no. 206508/Z/17/Z). J.C. receives support from the Medical Research Council Newton Fund (#MR/P017568/1).

Background Strongyloides stercoralis infection presents with varying degrees of severity, but it often primarily involves the small bowel. In severe infection and cases of hyperinfection, ileus and small-bowel obstruction may prevent enteral absorption of anthelminthics such as ivermectin. At present there are no parenteral anthelminthics licensed for use in humans. Methods Here, we describe two cases of severe S. stercoralis infection treated with an unlicensed veterinary preparation of subcutaneous ivermectin, and we discuss the published reports of the use of this treatment elsewhere. Results Both patients were successfully treated with subcutaneous ivermectin, and both recovered completely. Conclusions Despite the limited published experience of parenteral ivermectin use, there is evidence that it may be a safe and effective treatment for severe strongyloidiasis. However, more data are needed to guide dosing schedules and monitoring for toxicity.

OBJECTIVE: To examine temporal and geographical trends, risk factors, and seasonality of imported vivax malaria in the United Kingdom to inform clinical advice and policy. DESIGN: Observational study. SETTING: National surveillance data from the UK Public Health England Malaria Reference Laboratory, data from the International Passenger Survey, and international climactic data. PARTICIPANTS: All confirmed and notified cases of malaria in the UK (n=50,187) from 1987 to 2013, focusing on 12,769 cases of vivax malaria. MAIN OUTCOME MEASURES: Mortality, sociodemographic details (age, UK region, country of birth and residence, and purpose of travel), destination, and latency (time between arrival in the UK and onset of symptoms). RESULTS: Of the malaria cases notified, 25.4% (n=12,769) were due to Plasmodium vivax, of which 78.6% were imported from India and Pakistan. Most affected patients (53.5%) had travelled to visit friends and relatives, and 11.1% occurred in tourists. Imported P vivax is concentrated in areas with large communities of south Asian heritage. Overall mortality was 7/12,725 (0.05%), but with no deaths in 9927 patients aged under 50 years. Restricting the analysis to those aged more than 50 years, mortality was 7/2798 (0.25%), increasing to 4/526 (0.76%) (adjusted odds ratio 32.0, 95% confidence interval 7.1 to 144.0, P70 years) are most at risk of dying; these groups should be targeted for advice before travelling. The risk of acquiring vivax malaria is year round but higher during summer monsoons, masked by latency. The latency of time to clinical presentation of imported vivax malaria in the UK is highly seasonal; seasonal latency has implications for pretravel advice but also for the control of malaria in India and Pakistan. A reduced incidence of vivax malaria in travellers may mean further areas of South Asia can be considered not to need malaria chemoprophylaxis.

‘General languor and debility, feebleness of the heart's action, irritability of the stomach, and a peculiar change of the colour of the skin’ – forms part of the original description of Addison's disease, primary hypoadrenalism.1 The condition represents a diagnostic challenge for the clinician, a non-specific pattern of presentation notoriously difficult to identify, that follows a potentially fatal course. Data on the occurrence of spontaneous bilateral adrenal haemorrhage is sparse though it is becoming increasingly recognized. The case and corresponding literature are discussed to highlight the risk factors and clinical presentation of the condition.

OBJECTIVES: To quantify geographical concentration of falciparum malaria cases in the UK at a hospital level. To assess potential delay-to-treatment associated with hub-and-spoke distribution of artesunate in severe cases. DESIGN: Observational study using national and hospital data. SETTING AND PARTICIPANTS: 3520 patients notified to the Malaria Reference Laboratory 2008-2010, 34 patients treated with intravenous artesunate from a tropical diseases centre 2002-2010. MAIN OUTCOME MEASURES: Geographical location of falciparum cases notified in the UK. Diagnosis-to-treatment times for intravenous artesunate. RESULTS: Eight centres accounted for 43.9% of the UK's total cases; notifications from 107 centres accounted for 10.2% of cases; 51.5% of hospitals seeing malaria notified 5 or fewer cases in 3 years. Centres that saw