Diagnostic performance of mycological tests for invasive pulmonary aspergillosis in non-haematological patients: protocol for a systematic review and meta-analysis.

INTRODUCTION: Increasing numbers of patients with non-haematological diseases are infected with invasive pulmonary aspergillosis (IPA), with a high mortality reported which is mainly due to delayed diagnosis. The diagnostic capability of mycological tests for IPA including galactomannan test, (1,3)-ß-D-glucan test, lateral flow assay, lateral flow device and PCR for the non-haematological patients remains unknown. This protocol aims to conduct a systematic review and meta-analysis of the diagnostic performance of mycological tests to facilitate the early diagnosis and treatments of IPA in non-haematological diseases. METHODS AND ANALYSIS: Database including PubMed, CENTRAL and EMBASE will be searched from 2002 until the publication of results. Cohort or cross-sectional studies that assessing the diagnostic capability of mycological tests for IPA in patients with non-haematological diseases will be included. The true-positive, false-positive, true-negative and false-negative of each test will be extracted and pooled in bivariate random-effects model, by which the sensitivity and specificity will be calculated with 95% CI. The second outcomes will include positive (negative) likelihood ratio, area under the receiver operating characteristic curve and diagnostic OR will also be computed in the bivariate model. When applicable, subgroup analysis will be performed with several prespecified covariates to explore potential sources of heterogeneity. Factors that may impact the diagnostic effects of mycological tests will be examined by sensitivity analysis. The risk of bias will be appraised by the Quality Assessment tool for Diagnostic Accuracy Studies (QUADAS-2). ETHICS AND DISSEMINATION: This protocol is not involved with ethics approval, and the results will be peer-reviewed and disseminated on a recognised journal. PROSPERO REGISTRATION NUMBER: CRD42021241820.

Producing molecular biology reagents without purification.

We recently developed 'cellular' reagents-lyophilized bacteria overexpressing proteins of interest-that can replace commercial pure enzymes in typical diagnostic and molecular biology reactions. To make cellular reagent technology widely accessible and amenable to local production with minimal instrumentation, we now report a significantly simplified method for preparing cellular reagents that requires only a common bacterial incubator to grow and subsequently dry enzyme-expressing bacteria at 37°C with the aid of inexpensive chemical desiccants. We demonstrate application of such dried cellular reagents in common molecular and synthetic biology processes, such as PCR, qPCR, reverse transcription, isothermal amplification, and Golden Gate DNA assembly, in building easy-to-use testing kits, and in rapid reagent production for meeting extraordinary diagnostic demands such as those being faced in the ongoing SARS-CoV-2 pandemic. Furthermore, we demonstrate feasibility of local production by successfully implementing this minimized procedure and preparing cellular reagents in several countries, including the United Kingdom, Cameroon, and Ghana. Our results demonstrate possibilities for readily scalable local and distributed reagent production, and further instantiate the opportunities available via synthetic biology in general.

RT-PCR/MALDI-TOF mass spectrometry-based detection of SARS-CoV-2 in saliva specimens.

As severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infections continue, there is a substantial need for cost-effective and large-scale testing that utilizes specimens that can be readily collected from both symptomatic and asymptomatic individuals in various community settings. Although multiple diagnostic methods utilize nasopharyngeal specimens, saliva specimens represent an attractive alternative as they can rapidly and safely be collected from different populations. While saliva has been described as an acceptable clinical matrix for the detection of SARS-CoV-2, evaluations of analytic performance across platforms for this specimen type are limited. Here, we used a novel sensitive RT-PCR/MALDI-TOF mass spectrometry-based assay (Agena MassARRAY®) to detect SARS-CoV-2 in saliva specimens. The platform demonstrated high diagnostic sensitivity and specificity when compared to matched patient upper respiratory specimens. We also evaluated the analytical sensitivity of the platform and determined the limit of detection of the assay to be 1562.5 copies/ml. Furthermore, across the five individual target components of this assay, there was a range in analytic sensitivities for each target with the N2 target being the most sensitive. Overall, this system also demonstrated comparable performance when compared to the detection of SARS-CoV-2 RNA in saliva by the cobas® 6800/8800 SARS-CoV-2 real-time RT-PCR Test (Roche). Together, we demonstrate that saliva represents an appropriate matrix for SARS-CoV-2 detection on the novel Agena system as well as on a conventional real-time RT-PCR assay. We conclude that the MassARRAY® system is a sensitive and reliable platform for SARS-CoV-2 detection in saliva, offering scalable throughput in a large variety of clinical laboratory settings.

Comparison of saliva with oral and nasopharyngeal swabs for SARS-CoV-2 detection on various commercial and laboratory-developed assays.

The accurate laboratory detection of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a crucial element in the fight against coronavirus disease 2019 (COVID-19). Reverse transcription-polymerase chain reaction testing on combined oral and nasopharyngeal swab (ONPS) suffers from several limitations, including the need for qualified personnel, the discomfort caused by invasive nasopharyngeal sample collection, and the possibility of swab and transport media shortage. Testing on saliva would represent an advancement. The aim of this study was to compare the concordance between saliva samples and ONPS for the detection of SARS-CoV-2 on various commercial and laboratory-developed tests (LDT). Individuals were recruited from eight institutions in Quebec, Canada, if they had SARS-CoV-2 RNA detected on a recently collected ONPS, and accepted to provide another ONPS, paired with saliva. Assays available in the different laboratories (Abbott RealTime SARS-CoV-2, Cobas® SARS-CoV-2, Simplexa™ COVID-19 Direct, Allplex™ 2019-nCoV, RIDA®GENE SARS-CoV-2, and an LDT preceded by three different extraction methods) were used to determine the concordance between saliva and ONPS results. Overall, 320 tests were run from a total of 125 saliva and ONPS sample pairs. All assays yielded similar sensitivity when saliva was compared to ONPS, with the exception of one LDT (67% vs. 93%). The mean difference in cycle threshold (∆C t ) was generally (but not significantly) in favor of the ONPS for all nucleic acid amplification tests. The maximum mean ∆​​​​​C t was 2.0, while individual ∆C t varied importantly from -17.5 to 12.4. Saliva seems to be associated with sensitivity similar to ONPS for the detection of SARS-CoV-2 by various assays.

The value of anal swab RT-PCR for COVID-19 diagnosis in adult Indonesian patients.

OBJECTIVE: This study will test the performance of the anal swab PCR test when compared with the nasopharyngeal swab PCR test as a diagnostic tool for COVID-19. DESIGN: An observational descriptive study which included hospitalised suspected, or probable cases of hopitalised COVID-19 patients, conducted in Dr. Cipto Mangunkusumo National Hospital, Ciputra Hospital, Mitra Keluarga Depok Hospital and Mitra Keluarga Kelapa Gading Hospital, Indonesia. Epidemiological, clinical, laboratory and radiology data were obtained. Nasopharyngeal and anal swabs specimens were collected for SARS-CoV-2 RNA detection. RESULTS: We analysed 136 subjects as part of this study. The clinical spectrum of COVID-19 manifesation in this study was typical of hospitalised patients, with 25% classified as mild cases, 14.7% in severe condition and 12.5% of subjects classified as having acute respiratory distress syndrome. When compared with nasopharyngeal swab as the standard specimen for reverse transcription polymerase chain reaction (RT-PCR) detection of SARS-CoV-2 antigen, the sensitivity and specificity of the anal swab was 36.7% and 93.8%, respectively. The positive and negative predictive value were 97.8% and 16.5 %, respectively. The performance of the anal swab remained similar when only the subgroup of patients with gastrointestinal symptoms (n=92, 67.6%) was analysed (sensitivity 40% and specificity 91.7%). Out of all the subjects included in analysis, 67.6% had gastrointestinal symptoms. Similarly, 73.3% of patients in the anal swab-positive group had gastrointestinal symptoms. The two most common gastrointestinal symptoms in the subjects' population were nausea and anorexia. CONCLUSION: Anal swab specimen has low sensitivity (36.7%) but high specificity (93.8%) for detecting SARS-CoV-2 antigen by RT-PCR. Only one additional positive result was found by anal swab among the nasopharyngeal swab-negative group. Anal swab may not be needed as an additional test at the beginning of a patient's diagnostic investigation and nasopharyngeal swab RT-PCR remains as the standard diagnostic test for COVID-19.

Routine laboratory testing to determine if a patient has COVID-19.

BACKGROUND: Specific diagnostic tests to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and resulting COVID-19 disease are not always available and take time to obtain results. Routine laboratory markers such as white blood cell count, measures of anticoagulation, C-reactive protein (CRP) and procalcitonin, are used to assess the clinical status of a patient. These laboratory tests may be useful for the triage of people with potential COVID-19 to prioritize them for different levels of treatment, especially in situations where time and resources are limited. OBJECTIVES: To assess the diagnostic accuracy of routine laboratory testing as a triage test to determine if a person has COVID-19. SEARCH METHODS: On 4 May 2020 we undertook electronic searches in the Cochrane COVID-19 Study Register and the COVID-19 Living Evidence Database from the University of Bern, which is updated daily with published articles from PubMed and Embase and with preprints from medRxiv and bioRxiv. In addition, we checked repositories of COVID-19 publications. We did not apply any language restrictions. SELECTION CRITERIA: We included both case-control designs and consecutive series of patients that assessed the diagnostic accuracy of routine laboratory testing as a triage test to determine if a person has COVID-19. The reference standard could be reverse transcriptase polymerase chain reaction (RT-PCR) alone; RT-PCR plus clinical expertise or and imaging; repeated RT-PCR several days apart or from different samples; WHO and other case definitions; and any other reference standard used by the study authors. DATA COLLECTION AND ANALYSIS: Two review authors independently extracted data from each included study. They also assessed the methodological quality of the studies, using QUADAS-2. We used the 'NLMIXED' procedure in SAS 9.4 for the hierarchical summary receiver operating characteristic (HSROC) meta-analyses of tests for which we included four or more studies. To facilitate interpretation of results, for each meta-analysis we estimated summary sensitivity at the points on the SROC curve that corresponded to the median and interquartile range boundaries of specificities in the included studies. MAIN RESULTS: We included 21 studies in this review, including 14,126 COVID-19 patients and 56,585 non-COVID-19 patients in total. Studies evaluated a total of 67 different laboratory tests. Although we were interested in the diagnotic accuracy of routine tests for COVID-19, the included studies used detection of SARS-CoV-2 infection through RT-PCR as reference standard. There was considerable heterogeneity between tests, threshold values and the settings in which they were applied. For some tests a positive result was defined as a decrease compared to normal vaues, for other tests a positive result was defined as an increase, and for some tests both increase and decrease may have indicated test positivity. None of the studies had either low risk of bias on all domains or low concerns for applicability for all domains. Only three of the tests evaluated had a summary sensitivity and specificity over 50%. These were: increase in interleukin-6, increase in C-reactive protein and lymphocyte count decrease. Blood count Eleven studies evaluated a decrease in white blood cell count, with a median specificity of 93% and a summary sensitivity of 25% (95% CI 8.0% to 27%; very low-certainty evidence). The 15 studies that evaluated an increase in white blood cell count had a lower median specificity and a lower corresponding sensitivity. Four studies evaluated a decrease in neutrophil count. Their median specificity was 93%, corresponding to a summary sensitivity of 10% (95% CI 1.0% to 56%; low-certainty evidence). The 11 studies that evaluated an increase in neutrophil count had a lower median specificity and a lower corresponding sensitivity. The summary sensitivity of an increase in neutrophil percentage (4 studies) was 59% (95% CI 1.0% to 100%) at median specificity (38%; very low-certainty evidence). The summary sensitivity of an increase in monocyte count (4 studies) was 13% (95% CI 6.0% to 26%) at median specificity (73%; very low-certainty evidence). The summary sensitivity of a decrease in lymphocyte count (13 studies) was 64% (95% CI 28% to 89%) at median specificity (53%; low-certainty evidence). Four studies that evaluated a decrease in lymphocyte percentage showed a lower median specificity and lower corresponding sensitivity. The summary sensitivity of a decrease in platelets (4 studies) was 19% (95% CI 10% to 32%) at median specificity (88%; low-certainty evidence). Liver function tests The summary sensitivity of an increase in alanine aminotransferase (9 studies) was 12% (95% CI 3% to 34%) at median specificity (92%; low-certainty evidence). The summary sensitivity of an increase in aspartate aminotransferase (7 studies) was 29% (95% CI 17% to 45%) at median specificity (81%) (low-certainty evidence). The summary sensitivity of a decrease in albumin (4 studies) was 21% (95% CI 3% to 67%) at median specificity (66%; low-certainty evidence). The summary sensitivity of an increase in total bilirubin (4 studies) was 12% (95% CI 3.0% to 34%) at median specificity (92%; very low-certainty evidence). Markers of inflammation The summary sensitivity of an increase in CRP (14 studies) was 66% (95% CI 55% to 75%) at median specificity (44%; very low-certainty evidence). The summary sensitivity of an increase in procalcitonin (6 studies) was 3% (95% CI 1% to 19%) at median specificity (86%; very low-certainty evidence). The summary sensitivity of an increase in IL-6 (four studies) was 73% (95% CI 36% to 93%) at median specificity (58%) (very low-certainty evidence). Other biomarkers The summary sensitivity of an increase in creatine kinase (5 studies) was 11% (95% CI 6% to 19%) at median specificity (94%) (low-certainty evidence). The summary sensitivity of an increase in serum creatinine (four studies) was 7% (95% CI 1% to 37%) at median specificity (91%; low-certainty evidence). The summary sensitivity of an increase in lactate dehydrogenase (4 studies) was 25% (95% CI 15% to 38%) at median specificity (72%; very low-certainty evidence). AUTHORS' CONCLUSIONS: Although these tests give an indication about the general health status of patients and some tests may be specific indicators for inflammatory processes, none of the tests we investigated are useful for accurately ruling in or ruling out COVID-19 on their own. Studies were done in specific hospitalized populations, and future studies should consider non-hospital settings to evaluate how these tests would perform in people with milder symptoms.

Evaluating 10 Commercially Available SARS-CoV-2 Rapid Serological Tests by Use of the STARD (Standards for Reporting of Diagnostic Accuracy Studies) Method.

Numerous severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) rapid serological tests have been developed, but their accuracy has usually been assessed using very few samples, and rigorous comparisons between these tests are scarce. In this study, we evaluated and compared 10 commercially available SARS-CoV-2 rapid serological tests using the STARD (Standards for Reporting of Diagnostic Accuracy Studies) methodology. Two hundred fifty serum samples from 159 PCR-confirmed SARS-CoV-2 patients (collected 0 to 32 days after the onset of symptoms) were tested with rapid serological tests. Control serum samples (n = 254) were retrieved from pre-coronavirus disease (COVID) periods from patients with other coronavirus infections (n = 11), positivity for rheumatoid factors (n = 3), IgG/IgM hyperglobulinemia (n = 9), malaria (n = 5), or no documented viral infection (n = 226). All samples were tested using rapid lateral flow immunoassays (LFIAs) from 10 manufacturers. Only four tests achieved ≥98% specificity, with the specificities ranging from 75.7% to 99.2%. The sensitivities varied by the day of sample collection after the onset of symptoms, from 31.7% to 55.4% (days 0 to 9), 65.9% to 92.9% (days 10 to 14), and 81.0% to 95.2% (>14 days). Only three of the tests evaluated met French health authorities' thresholds for SARS-CoV-2 serological tests (≥90% sensitivity and ≥98% specificity). Overall, the performances varied greatly between tests, with only one-third meeting acceptable specificity and sensitivity thresholds. Knowledge of the analytical performances of these tests will allow clinicians and, most importantly, laboratorians to use them with more confidence; could help determine the general population's immunological status; and may help diagnose some patients with false-negative real-time reverse transcription-PCR (RT-PCR) results.

Laboratory Tests and Outcome for Patients with Coronavirus Disease 2019: A Systematic Review and Meta-Analysis.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 causes coronavirus disease 2019 (COVID-19) and poses substantial challenges for healthcare systems. With a vastly expanding number of publications on COVID-19, clinicians need evidence synthesis to produce guidance for handling patients with COVID-19. In this systematic review and meta-analysis, we examine which routine laboratory tests are associated with severe COVID-19 disease. CONTENT: PubMed (Medline), Scopus, and Web of Science were searched until March 22, 2020, for studies on COVID-19. Eligible studies were original articles reporting on laboratory tests and outcome of patients with COVID-19. Data were synthesized, and we conducted random-effects meta-analysis, and determined mean difference (MD) and standard mean difference at the biomarker level for disease severity. Risk of bias and applicability concerns were evaluated using the Quality Assessment of Diagnostic Accuracy Studies-2. SUMMARY: 45 studies were included, of which 21 publications were used for the meta-analysis. Studies were heterogeneous but had low risk of bias and applicability concern in terms of patient selection and reference standard. Severe disease was associated with higher white blood cell count (MD, 1.28 ×109/L), neutrophil count (MD, 1.49 ×109/L), C-reactive protein (MD, 49.2 mg/L), lactate dehydrogenase (MD, 196 U/L), D-dimer (standardized MD, 0.58), and aspartate aminotransferase (MD, 8.5 U/L); all p < 0.001. Furthermore, low lymphocyte count (MD -0.32 × 109/L), platelet count (MD -22.4 × 109/L), and hemoglobin (MD, -4.1 g/L); all p < 0.001 were also associated with severe disease. In conclusion, several routine laboratory tests are associated with disease severity in COVID-19.

Dynamics of SARS-CoV-2 RT-PCR positivity and seroprevalence among high-risk healthcare workers and hospital staff.

Staff working in units that were highly exposed to coronavirus disease 2019 were invited to participate in a 6-month study on the carriage and seroprevalence of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). The results from visits on Day 1 and Day 15 show that 41 cases of SARS-CoV-2 infection were confirmed by reverse transcriptase polymerase chain reaction and/or serology in 326 participants (overall infection rate 12.6%). The presence of comorbidities or symptoms at the time of sample collection was a risk factor for infection, but working as a physician/nurse was not a risk factor. Universal screening in high-risk units, irrespective of symptoms, allowed the identification of asymptomatic and potentially contagious infected workers, enabling them to self-isolate for 7 days.

Diagnostic impact of bedside chest X-ray features of 2019 novel coronavirus in the routine admission at the emergency department: case series from Lombardy region.

PURPOSE: To evaluate the diagnostic accuracy and the imaging features of routine admission chest X-ray in patients suspected for novel Coronavirus 2019 (SARS-CoV-2) infection. METHOD: We retrospectively evaluated clinical and X-ray features in all patients referred to the emergency department for suspected SARS-CoV-2 infection between March 1st and March 13th. A single radiologist with more than 15 years of experience in chest-imaging evaluated the presence and extent of alveolar opacities, reticulations, and/or pleural effusion. The percentage of lung involvement (range <25 % to 75-100 %) was also calculated. We stratified patients in groups according to the time interval between symptoms onset and X-ray imaging (≤ 5 and > 5 days) and according to age (≤ 50 and > 50 years old). RESULTS: A total of 518 patients were enrolled. Overall 314 patients had negative and 204 had positive RT-PCR results. Lung lesions in patients with SARS-Cov2 pneumonia primarily manifested as alveolar and interstitial opacities and were mainly bilateral (60.8 %). Lung abnormalities were more frequent and more severe by symptom duration and by increasing age. The sensitivity and specificity of chest X-ray at admission in the overall cohort were 57 % (95 % CI = 47-67) and 89 % (83-94), respectively. Sensitivity was higher for patients with symptom onset > 5 days compared to ≤ 5 days (76 % [62-87] vs 37 % [24-52]) and in patients > 50 years old compared to ≤ 50 years (59 % [48-69] vs 47 % [23-72]), at the expense of a slightly lower specificity (68 % [45-86] and 82 % [73-89], respectively). CONCLUSIONS: Overall chest X-ray sensitivity for SARS-CoV-2 pneumonia was 57 %. Sensitivity was higher when symptoms had started more than 5 days before, at the expense of lesser specificity, while slightly higher in older patients in comparison to younger ones.

There is no routine head and neck exam during the COVID-19 pandemic.

Head and neck examinations are commonly performed by all physicians. In the era of the COVID-19 pandemic caused by the SARS-CoV-2 virus, which has a high viral load in the upper airways, these examinations and procedures of the upper aerodigestive tract must be approached with caution. Based on experience and evidence from SARS-CoV-1 and early experience with SARS-CoV-2, we provide our perspective and guidance on mitigating transmission risk during head and neck examination, upper airway endoscopy, and head and neck mucosal surgery including tracheostomy.

Recurrence of positive SARS-CoV-2 RNA in COVID-19: A case report.

The ongoing outbreak of COVID-19 that began in Wuhan, China, has constituted a Public Health Emergency of International Concern, with cases confirmed in multiple countries. Currently, patients are the primary source of infection. We report a confirmed case of COVID-19 whose oropharyngeal swab test of SARS-CoV-2 RNA turned positive in convalescence. This case highlights the importance of active surveillance of SARS-CoV-2 RNA for infectivity assessment.