Implications of multiple freeze-thawing on respiratory samples for culture-independent analyses

© 2014 . Background: Best practice when performing culture-independent microbiological analysis of sputum samples involves their rapid freezing and storage at -80 °C. However, accessing biobanked collections can mean that material has been passed through repeated freeze-thaw cycles. The aim of this study was to determine the impact of these cycles on microbial community profiles. Methods: Sputum was collected from eight adults with cystic fibrosis, and each sample was subjected to six freeze-thaw cycles. Following each cycle, an aliquot was removed and treated with propidium monoazide (PMA) prior to DNA extraction and 16S rRNA gene pyrosequencing. Results: The impact of freeze-thaw cycles was greatest on rare members of the microbiota, with variation beyond that detected with within-sample repeat analysis observed after three cycles. Conclusion: Four or more freeze thaw cycles result in a significant distortion of microbiota profiles from CF sputum

Immunosuppressive therapy after solid organ transplantation and the gut microbiota: bidirectional interactions with clinical consequences

International audienceOur understanding of the involvement of the gut microbiota (GM) in human health has expanded exponentially over the last few decades, particularly in the fields of metabolism, inflammation, and immunology. Immunosuppressive treatment (IST) prescribed to solid organ transplant (SOT) recipients produces GM changes that affect these different processes. This review aims at describing the current knowledge of how IST changes the GM. Overall, SOT followed by IST results in persistent changes in the GM, with a consistent increase in proteobacteria including opportunistic pathobionts. In mice, Tacrolimus induces dysbiosis and metabolic disorders, and alters the intestinal barrier. The transfer of the GM from Tacrolimus-treated hosts confers immunosuppressive properties, suggesting a contributory role for the GM in this drug's efficacy. Steroids induce dysbiosis and intestinal barrier alterations, and also seem to depend partly on the GM for their immunosuppressive and metabolic effects. Mycophenolate Mofetil, frequently responsible for digestive side effects such as diarrhea and colitis, is associated with pro-inflammatory dysbiosis and increased endotoxemia. Alemtuzumab, m-TOR inhibitors, and belatacept have shown more marginal impact on the GM. Most of these observations are descriptive. Future studies should explore the underlying mechanism of IST-induced dysbiosis in order to better understand their efficacy and safety characteristic

Performance and ease of use of a molecular point-of-care test for influenza A/B and RSV in patients presenting to primary care

Annual influenza epidemics cause substantial morbidity and mortality, and the majority of patients with influenza-like illness present to primary care. Point-of-care influenza tests could support treatment decisions. It is critical to establish analytic performance of these platforms in real-life patient samples before uptake can be considered. We aimed to assess the analytical performance and ease of use of the cobas® Liat® PCR POCT in detecting influenza A/B and RSV in samples collected from patients with influenza-like illness in primary care. Sensitivity and specificity of the cobas® Liat® POCT are calculated in comparison with a commercial laboratory-based PCR test (Fast-Track Respiratory Pathogens 21 Plus kit (Fast-Track Diagnostics)). Samples with discordant results were analysed additionally by the RespiFinder 2Smart (PathoFinder) using an Extended Gold Standard (EGS). Acceptability was scored on a five-point Likert scale as well as a failure mode analysis of the cobas® Liat® POCT was performed. Nasal and oropharyngeal swabs were obtained from 140 children and nasopharyngeal swabs from 604 adults (744 patients). The cobas® Liat® POCT had a sensitivity and specificity of 100% (95% CI 99-100%) and 98.1% (95%CI 96.3-99%) for influenza A, 100% (95% CI 97.7-100%) and 99.7% (95%CI 98.7-99.9%) for influenza B and 100% (95% CI 87.1-100%) and 99.4% (95%CI 98.6-99.8%) for RSV, respectively. According to trained lab technicians, the cobas® Liat® POCT was considered easy-to-use, with a fast turn-around-time. Cobas® Liat® POCT is a promising decentralised test platform for influenza A/B and RSV in primary care as it provides fairly rapid results with excellent analytic performance. Point-of-care influenza tests could support treatment decisions in primary care. Cobas® Liat® POCT is a promising decentralised test platform for influenza A/B and RSV in primary care as it provides fairly rapid results with excellent analytic performance.status: publishe

Disease course of lower respiratory tract infection with a bacterial cause

PURPOSE Bacterial pathogens are assumed to cause an illness course different from that of nonbacterial causes of acute cough, but evidence is lacking. We evaluated the disease course of lower respiratory tract infection (LRTI) with a bacterial cause in adults with acute cough. METHODS We conducted a secondary analysis of a multicenter European trial in which 2,061 adults with acute cough (28 days’ duration or less) were recruited from primary care and randomized to amoxicillin or placebo. For this analysis only patients in the placebo group (n = 1,021) were included, reflecting the natural course of disease. Standardized microbiological and serological analyses were performed at baseline to define a bacterial cause. All patients recorded symptoms in a diary for 4 weeks. The disease course between those with and without a bacterial cause was compared by symptom severity in days 2 to 4, duration of symptoms rated moderately bad or worse, and a return consultation. RESULTS Of 1,021 eligible patients, 187 were excluded for missing diary records, leaving 834 patients, of whom 162 had bacterial LRTI. Patients with bacterial LRTI had worse symptoms at day 2 to 4 after the first office visit (P = .014) and returned more often for a second consultation, 27% vs 17%, than those without bacterial LRTI (P = .004). Resolution of symptoms rated moderately bad or worse did not differ (P = .375). CONCLUSIONS Patients with acute bacterial LRTI have a slightly worse course of disease when compared with those without an identified bacterial cause, but the relevance of this difference is not meaningful

Clinical diagnosis of SARS-CoV-2 infection: An observational study of respiratory tract infection in primary care in the early phase of the pandemic

AbstractBackground Early in the COVID-19 pandemic, GPs had to distinguish SARS-CoV-2 from other aetiologies in patients presenting with respiratory tract infection (RTI) symptoms on clinical grounds and adapt management accordingly.Objectives To test the diagnostic accuracy of GPs’ clinical diagnosis of a SARS-CoV-2 infection in a period when COVID-19 was a new disease. To describe GPs’ management of patients presenting with RTI for whom no confirmed diagnosis was available. To investigate associations between patient and clinical features with a SARS-CoV-2 infection.Methods In April 2020–March 2021, 876 patients (9 countries) were recruited when they contacted their GP with symptoms of an RTI of unknown aetiology. A swab was taken at baseline for later analysis. Aetiology (PCR), diagnostic accuracy of GPs’ clinical SARS-CoV-2 diagnosis, and patient management were explored. Factors related to SARS-CoV-2 infection were determined by logistic regression modelling.Results GPs suspected SARS-CoV-2 in 53% of patients whereas 27% of patients tested positive for SARS-CoV-2. True-positive patients (23%) were more intensively managed for follow-up, antiviral prescribing and advice than true-negatives (42%). False negatives (5%) were under-advised, particularly for social distancing and isolation. Older age (OR: 1.02 (1.01–1.03)), male sex (OR: 1.68 (1.16–2.41)), loss of taste/smell (OR: 5.8 (3.7–9)), fever (OR: 1.9 (1.3–2.8)), muscle aches (OR: 2.1 (1.5–3)), and a known risk factor for COVID-19 (travel, health care worker, contact with proven case; OR: 2.7 (1.8–4)) were predictive of SARS-CoV-2 infection. Absence of loss of taste/smell, fever, muscle aches and a known risk factor for COVID-19 correctly excluded SARS-CoV-2 in 92.3% of patients, whereas presence of 3, or 4 of these variables correctly classified SARS-CoV-2 in 57.7% and 87.1%.Conclusion Correct clinical diagnosis of SARS-CoV-2 infection, without POC-testing available, appeared to be complicated

Aetiology of lower respiratory tract infection in adults in primary care: A prospective study in 11 European Countries

Objectives: To describe the role of bacteria (including bacterial resistance), viruses (including those recently described), and mixed bacterial-viral infections in adults presenting to primary care with lower respiratory tract infection (LRTI). Methods: We enrolled 3104 adults with LRTI, 141 (4.5%) of whom had community-acquired pneumonia (CAP), and 2985 matched controls in a prospective study in 16 primary care networks in Europe, and followed patients up at 28-35 days. We detected S. pneumoniae and H. influenzae and assessed susceptibility, atypical bacteria and viruses. Results: A potential pathogen was detected in 1844 (59%) (in 350 (11%) bacterial pathogens only, in 1190 (38%) viral pathogens only, and in 304 (10%) both bacterial and viral pathogens). The most common bacterial pathogens isolated were S. pneumoniae (5.5% overall, 9.2% in CAP patients) and H. influenzae (5.4% overall, 14.2% in CAP patients). <1% of S. pneumoniae were highly resistant to penicillin and 12.6% of H. influenzae were beta-lactamase positive. The most common viral pathogens detected were human rhinovirus (HRV; 20.1%), influenza viruses (FLU; 9.9%), and human coronavirus (HCoV; 7.4%). FLU, human parainfluenzaviruses and human respiratory syncytial virus as well as HRV, HCoV, human metapneumovirus were detected significantly more frequently in LRTI patients than in controls. Conclusions: A bacterial pathogen is identified in approximately one in five adult patients with LRTI in primary care, and a viral pathogen in just under half, with mixed infections in one in ten. Penicillin resistant pneumococci and beta-lactamase producing H. influenzae are uncommon. These new findings support a restrictive approach to antibiotic prescribing for LRTI and the use of first-line, narrow spectrum agents in primary care

COVID-19 in Patients on Maintenance Dialysis in the Paris Region

International audienceIntroduction: Coronavirus disease 2019 (COVID-19) represents a serious threat to patients on maintenance dialysis. The clinical setting, mortality rate, and prognostic factors in these patients have not been well established. Methods: We included all dialyzed patients with COVID-19 referred to our dialysis center between March 11 and April 11, 2020. Data were obtained through the review of the medical records and were censored at the time of data cutoff, on May 11, 2020. Results: Forty-four patients on maintenance dialysis with COVID-19 were referred to our dialysis unit during the COVID-19 epidemic. Median age was 61 years (interquartile range [IQR]: 51.5–72.5); 65.9% were men. Comorbidities included hypertension (97.7%), diabetes mellitus (50%), and chronic cardiac (38.6%) and respiratory (27.3%) diseases. Initial symptoms were fever (79.5%), shortness of breath (29.5%), cough (43.2%), and diarrhea (13.6%). Three profiles of severity were distinguished based on the World Health Organization (WHO) progression scale. Forty-one (93.2%) were hospitalized and only 3 were maintained on outpatient hemodialysis. Thirty-three (75%) patients required oxygen therapy, including 15 (45.5%) who were referred to the intensive care unit. Overall, 27.3% of patients died, and 58.5% were discharged from hospital, including only 2 (13.3%) of those admitted to the intensive care unit. By multivariate analysis, cough, thrombopenia 175 mg/l were significantly associated with death. Conclusion: A major outbreak of COVID-19 occurred in the Paris region, and spread among dialyzed patients. Our study underscores the severity of COVID-19 in these patients and identified prognostic markers