Background Tuberculosis remains a global health challenge, with early diagnosis key to its reduction. Face-mask sampling detects exhaled Mycobacterium tuberculosis. We aimed to investigate bacillary output from patients with pulmonary tuberculosis and to assess the potential of face-mask sampling as a diagnostic method in active casefinding. MethodsWe did a 24-h longitudinal study in patients from three hospitals in Pretoria, South Africa, with microbiologically confirmed pulmonary tuberculosis. Patients underwent 1 h of face-mask sampling eight times over a 24-h period, with contemporaneous sputum sampling. M tuberculosis was detected by quantitative PCR. We also did an active case-finding pilot study in inhabitants of an informal settlement near Pretoria. We enrolled individuals with symptoms of tuberculosis on the WHO screening questionnaire. Participants provided sputum and face-mask samples that were tested with the molecular assay Xpert MTB/RIF Ultra. Sputum-negative and face-mask-positive individuals were followed up prospectively for 20 weeks by bronchoscopy, PET-CT, and further sputum analysis to validate the diagnosis.
Background Human to human transmission of SARS-CoV-2 is driven by the respiratory route but little is known about the pattern and quantity of virus output from exhaled breath. We have previously shown that face-mask sampling (FMS) can detect exhaled tubercle bacilli and have adapted its use to quantify exhaled SARS-CoV-2 RNA in patients admitted to hospital with Coronavirus Disease-2019 (COVID-19). Methods Between May and December 2020, we took two concomitant FMS and nasopharyngeal samples (NPS) over two days, starting within 24 hours of a routine virus positive NPS in patients hospitalised with COVID-19, at University Hospitals of Leicester NHS Trust, UK. Participants were asked to wear a modified duckbilled facemask for 30 minutes, followed by a nasopharyngeal swab. Demographic, clinical, and radiological data, as well as International Severe Acute Respiratory and emerging Infections Consortium (ISARIC) mortality and deterioration scores were obtained. Exposed masks were processed by removal, dissolution and analysis of sampling matrix strips fixed within the mask by RT-qPCR. Viral genome copy numbers were determined and results classified as Negative; Low: ≤999 copies; Medium: 1,000-99,999 copies and High ≥ 100,000 copies per strip for FMS or per 100µl for NPS. Results 102 FMS and NPS were collected from 66 routinely positive patients; median age: 61 (IQR 49 - 77), of which FMS was positive in 38% of individuals and concomitant NPS was positive in 50%. Positive FMS viral loads varied over five orders of magnitude (<10-3.3 x 10 6 genome copies/strip); 21 (32%) patients were asymptomatic at the time of sampling. High FMS viral load was associated with respiratory symptoms at time of sampling and shorter interval between sampling and symptom onset (FMS High: median (IQR) 2 days (2-3) vs FMS Negative: 7 days (7-10), p =0.002). On multivariable linear regression analysis, higher FMS viral loads were associated with higher ISARIC mortality (Medium FMS vs Negative FMS gave an adjusted coefficient of 15.7, 95% CI 3.7-27.7, p =0.01) and deterioration scores (High FMS vs Negative FMS gave an adjusted coefficient of 37.6, 95% CI 14.0 to 61.3, p =0.002), while NPS viral loads showed no significant association. Conclusion We demonstrate a simple and effective method for detecting and quantifying exhaled SARS-CoV-2 in hospitalised patients with COVID-19. Higher FMS viral loads were more likely to be associated with developing severe disease compared to NPS viral loads. Similar to NPS, FMS viral load was highest in early disease and in those with active respiratory symptoms, highlighting the potential role of FMS in understanding infectivity.
IntroductionThe degree to which bacteria in the human respiratory tract are aerosolised by individuals is not established. Building on our experience sampling bacteria exhaled by individuals with pulmonary tuberculosis using face masks, we hypothesised that patients with conditions frequently treated with antimicrobials, such as chronic obstructive pulmonary disease (COPD), might exhale significant numbers of bacteria carrying antimicrobial resistance (AMR) genes and that this may constitute a previously undefined risk for the transmission of AMR.MethodsFifteen-minute mask samples were taken from 13 patients with COPD (five paired with contemporaneous sputum samples) and 10 healthy controls. DNA was extracted from cell pellets derived from gelatine filters mounted within the mask. Quantitative PCR analyses directed to the AMR encoding genes: blaTEM (β-lactamase), ErmB (target methylation), mefA (macrolide efflux pump) and tetM (tetracycline ribosomal protection protein) and six additional targets were investigated. Positive signals above control samples were obtained for all the listed genes; however, background signals from the gelatine precluded analysis of the additional targets.Results9 patients with COPD (69%), aerosolised cells containing, in order of prevalence, mefA, tetM, ErmB and blaTEM, while three healthy controls (30%) gave weak positive signals including all targets except blaTEM. Maximum estimated copy numbers of AMR genes aerosolised per minute were mefA: 3010, tetM: 486, ErmB: 92 and blaTEM: 24. The profile of positive signals found in sputum was not concordant with that in aerosol in multiple instances.DiscussionWe identified aerosolised AMR genes in patients repeatedly exposed to antimicrobials and in healthy volunteers at lower frequencies and levels. The discrepancies between paired samples add weight to the view that sputum content does not define aerosol content. Mask sampling is a simple approach yielding samples from all subjects and information distinct from sputum analysis. Our results raise the possibility that patient-generated aerosols may be a significant means of AMR dissemination that should be assessed further and that consideration be given to related control measures.
Background Nasopharyngeal samples (NPS) are the mainstay of COVID-19 diagnosis. However, the extent to which assay signals relate to exhaled virus is unknown. We investigated the use of novel, non-invasive face-mask sampling (FMS) to detect exhaled SARS-CoV-2 RNA in two studies. Methods In an outbreak study (cohort 1), we performed FMS and NPS for 21 consecutive days after diagnosis on six healthcare workers who were screened positive for SARS-CoV-2. In a second hospitalised cohort (cohort 2), we performed FMS on 47 patients within 24 hours of a positive diagnosis. COVID-19 severity was graded according to WHO recommendations. Findings In cohort 1, SARS-COV-2 was detected by FMS in 10/40 (25%) samples (4/6 individuals), with no correlation between NPS and FMS RNA signals. All samples were negative by day 14 post diagnosis. Sustained FMS positivity with higher viral RNA signals showed a trend towards disease severity. In cohort 2, 19/47 (40%) individuals exhaled SARS-CoV-2 RNA extending over five orders of magnitude. FMS positive participants were older (positive: median age [IQR] 71 [61-84] vs negative: 61 [45-73], p=0.04) with more comorbidities (positive: 2 [1-3] vs negative: 1 [0-2], p<0.001) and have active cough (positive: 68% vs negative: 24%, p=0.003) and breathlessness (positive: 74% vs negative: 32%, p=0.005) during sampling, compared to FMS negative patients. Of five patients who were FMS positive and asymptomatic at time of sampling, two died of severe COVID-19 pneumonia within one month of follow up. Interpretation FMS detects exhaled SARS-COV-2, with stronger signals in those who develop severe disease.
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