Lower Respiratory Tract Samples Research Articles

The human microbiome and COVID-19: A systematic review.

Human microbiotas are communities of microorganisms living in symbiosis with humans. They play an important role in the host immune response to respiratory viral infection. However, evidence on the human microbiome and coronavirus disease (COVID-19) relationship is insufficient. The aim of this systematic literature review was to evaluate existing evidence on the association between the microbiome and COVID-19 in humans and summarize these data in the pandemic era. We conducted a systematic literature review on the association between the microbiome and COVID-19 in humans by searching PubMed, Embase, and the Cochrane Library, CINAHL, and Web of Science databases for articles in English published up to October 31, 2020. The results were analyzed qualitatively. This study is registered with PROSPERO (CRD42020195982). Of the 543 articles identified by searching databases, 16 in line with the research objectives were eligible for qualitative review: eight sampled the microbiome using stool, four using nasopharyngeal or throat swab, three using bronchoalveolar lavage fluid, and one using lung tissue. Fecal microbiome dysbiosis and increased opportunistic pathogens were reported in COVID-19 patients. Several studies suggested the dysbiosis in the lung microbiome of COVID-19 patients with an abundance of opportunistic pathogens using lower respiratory tract samples. The association between COVID-19 severity and the human microbiome remains uncertain. The human fecal and respiratory tract microbiome changed in COVID-19 patients with opportunistic pathogen abundance. Further research to elucidate the effect of alternation of the human microbiome in disease pathogenesis is warranted.

Modeling the early temporal dynamics of viral load in respiratory tract specimens of COVID-19 patients in Incheon, the Republic of Korea

ObjectiveTo investigate the duration and peak of severe acute respiratory syndrome coronavirus 2 shedding as infectivity markers for determining the isolation period. MethodsA total of 2,558 upper respiratory tract (URT) and lower respiratory tract (LRT) specimens from 138 patients with laboratory-confirmed coronavirus disease were analyzed. Measurements of sequential viral loads were aggregated using the cubic spline smoothing function of a generalized additive model. The time to negative conversion was compared between symptom groups using survival analysis. ResultsIn URT samples, viral RNA levels peaked on day 4 after symptom onset and rapidly decreased until day 10 for both E and RdRp genes, whereas those in LRT samples immediately peaked from symptom onset and decreased until days 15.6 and 15.0 for E and RdRp genes, respectively. Median (interquartile range) time to negative conversion was significantly longer in symptomatic (18.0 [13.0–25.0] days) patients than in asymptomatic (13.0 [9.5–17.5] days) patients. The more types of symptoms a patient had, the longer the time to negative conversion. ConclusionsThe viral load rapidly changes depending on the time after symptom onset; the viral shedding period may be longer with more clinical symptoms. Different isolation policies should be applied depending on disease severity.

High Levels of Neutrophil Extracellular Traps Persist in the Lower Respiratory Tract of Critically Ill Patients With Coronavirus Disease 2019.

Lower respiratory tract (LRT) disease induced by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can deteriorate to acute respiratory distress syndrome (ARDS). Because the release of neutrophil extracellular traps (NETs) is implicated in ARDS pathogenesis, we investigated the presence of NETs and correlates of pathogenesis in blood and LRT samples of critically ill patients with COVID-19. Plasma NET levels peaked early after intensive care unit admission and were correlated with the SARS-CoV-2 RNA load in sputum and levels of neutrophil-recruiting chemokines and inflammatory markers in plasma samples. The baseline plasma NET quantity was correlated with disease severity but was not associated with soluble markers of thrombosis or with development of thrombosis. High NET levels were present in LRT samples and persisted during the course of COVID-19, consistent with the detection of NETs in bronchi and alveolar spaces in lung tissue from deceased patient with COVID-19. Thus, NETs are produced and retained in the LRT of critically ill patients with COVID-19 and could contribute to SARS-CoV-2-induced ARDS disease.

The utility of paired upper and lower respiratory tract sampling for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) testing in patients with artificial airways.

Early in the coronavirus disease 2019 (COVID-19) pandemic, the CDC recommended collection of a lower respiratory tract (LRT) specimen for severe acute respiratory coronavirus virus 2 (SARS-CoV-2) testing in addition to the routinely recommended upper respiratory tract (URT) testing in mechanically ventilated patients. Significant operational challenges were noted at our institution using this approach. In this report, we describe our experience with routine collection of paired URT and LRT sample testing. Our results revealed a high concordance between the 2 sources, and that all children tested for SARS-CoV-2 were appropriately diagnosed with URT testing alone. There was no added benefit to LRT testing. Based on these findings, our institutional approach was therefore adjusted to sample the URT alone for most patients, with LRT sampling reserved for patients with ongoing clinical suspicion for SARS-CoV-2 after a negative URT test.

Detectable respiratory SARS-CoV-2 RNA is associated with low vitamin D levels and high social deprivation.

BackgroundAccumulating evidence links COVID‐19 incidence and outcomes with vitamin D status. We investigated if an interaction existed between vitamin D levels and social deprivation in those with and without COVID‐19 infection.MethodsUpper or lower respiratory tract samples from 104 patients were tested for SARS‐CoV‐2 RNA in accordance with Public Health England criteria (January‐May 2020) using RT‐PCR. The latest serum total 25‐hydroxyvitamin D(25‐OHD) levels, quantified by LC‐MS/MS, was obtained for each patient (September 2019‐April 2020). Index of Multiple Deprivation (IMD) was generated for each patient. Univariate and logistic regression analyses examined associations between age, gender, 25‐OHD, IMD score and SARS‐CoV‐2 result in the total cohort and subgroups.ResultsIn the total cohort, a positive SARS‐CoV‐2 test was significantly associated with lower 25‐OHD levels and higher IMD. A positive test was associated with higher IMD in the male subgroup and with lower 25‐OHD levels in those aged >72 years. Low 25‐OHD and IMD quintile 5 were separately associated with positive COVID‐19 outcome in the cohort. Patients in IMD quintile 5 with vitamin D levels ≤ 34.4 nmol/L were most likely to have a positive COVID‐19 outcome, even more so if aged >72 years (OR: 19.07, 95%CI: 1.71‐212.25; P = .016).ConclusionsIn this cohort, combined low vitamin D levels and higher social deprivation were most associated with COVID‐19 infection. In older age, this combination was even more significant. Our data support the recommendations for normalising vitamin D levels in those with deficient / insufficient levels and in groups at high risk for deficiency.

The Unyvero Hospital-Acquired pneumonia panel for diagnosis of secondary bacterial pneumonia in COVID-19 patients

The study was undertaken to evaluate the performance of Unyvero Hospitalized Pneumonia (HPN) panel application, a multiplex PCR-based method for the detection of bacterial pathogens from lower respiratory tract (LRT) samples, obtained from COVID-19 patients with suspected secondary hospital-acquired pneumonia. Residual LRT samples obtained from critically ill COVID-19 patients with predetermined microbiological culture results were tested using the Unyvero HPN Application. Performance evaluation of the HPN Application was carried out using the standard-of-care (SoC) microbiological culture findings as the reference method. Eighty-three LRT samples were used in the evaluation. The HPN Application had a full concordance with SoC findings in 59/83 (71%) samples. The new method detected additional bacterial species in 21 (25%) and failed at detecting a bacterial species present in lower respiratory culture in 3 (3.6%) samples. Overall the sensitivity, specificity, positive, and negative predictive values of the HPN Application were 95.1% (95%CI 96.5–98.3%), 98.3% (95% CI 97.5–98.9%), 71.6% (95% CI 61.0–80.3%), and 99.8% (95% CI 99.3–99.9%), respectively. In conclusion, the HPN Application demonstrated higher diagnostic yield in comparison with the culture and generated results within 5 h.

Surveillance of Coronavirus Disease 2019 (COVID-19) Testing in Clinical Laboratories in Korea

In response to the ongoing coronavirus disease 2019 (COVID-19) pandemic, an online laboratory surveillance system was established to monitor severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) real-time reverse transcription-PCR (rRT-PCR) testing capacities and results. SARS-CoV-2 rRT-PCR testing data were collected from 97 clinical laboratories, including 84 medical institutions and 13 independent clinical laboratories in Korea. We assessed the testing capacities to utilize SARS-CoV-2 rRT-PCR based on surveillance data obtained from February 7th to June 4th, 2020 and evaluated positive result characteristics according to the reagents used and sample types. A total of 1,890,319 SARS-CoV-2 rRT-PCR testing were performed, 2.3% of which were positive. Strong correlations were observed between the envelope (E) gene and RNA-dependent RNA polymerase (RdRp)/nucleocapsid (N) genes threshold cycle (Ct) values for each reagent. No statistically significant differences in gene Ct values were observed between the paired upper and lower respiratory tract samples, except in the N gene for nasopharyngeal swab and sputum samples. Our study showed that clinical laboratories in Korea have rapidly expanded their testing capacities in response to the COVID-19 outbreak, with a peak daily capacity of 34,193 tests. Rapid expansion in testing capacity is a critical component of the national response to the ongoing pandemic.

COVID-19 Microbiological and Radiological Diagnostics

The purpose of this paper is to present SARS-CoV-2 diagnostic methods and their optimal implementation. The main routine microbiological diagnostic tools are molecular and serological methods. Sensitivity and utilisation of molecular methods is the best at the disease onset, while serological methods are the best if used at least 5-7 days after disease onset. Positive molecular test result (real-time polymerase chain reaction; RT-PCR) must be interpreted in the clinical presentation context. A sampling site for molecular analysis is based on disease severity; upper and lower respiratory tract samples are used for mild and severe presentations, respectively. Virus cultivation is the only method of proving the existence of a viable virus in a tissue sample, but due to the method complexity, it is not a part of a routine process. Ground glass opacities with or without reticular pattern and/or consolidation are typical findings for COVID-19 pneumonia. Multi-slice computerized tomography (MSCT) is a superior radiological method for performing X-ray of the chests.

The Human Microbiome and COVID-19: A Systematic Review

Background: Human microbiotas are community of microorganisms, living in symbiosis with humans. They play an important role in the host immune response to respiratory viral infection. However, evidence on the human microbiome and coronavirus disease (COVID-19) relationship is insufficient. This systematic literature review aimed to evaluate existing evidence on the association between the microbiome and COVID-19 in humans and summarize this data in the pandemic era.Methods: We conducted a systematic literature review on the association between the microbiome and COVID-19 in humans by searching PubMed, Embase, and the Cochrane Library, CINAHL, and Web of Science databases for articles in English published up to October 31, 2020. The results were analyzed using qualitative analysis of evidence. This study is registered with PROSPERO (CRD42020195982).Findings: Of the 543 articles identified by searching databases, 16 in line with the research objectives were eligible for qualitative review: eight sampled the microbiome using stool, four using nasopharyngeal or throat swab, three using bronchoalveolar lavage fluid, and one using lung tissue. Fecal microbiome dysbiosis and increased opportunistic pathogens in COVID-19 patients were reported. Several studies suggested the dysbiosis in the lung microbiome of COVID-19 patients with abundance of opportunistic pathogens using lower respiratory tract samples. The association between COVID-19 severity and the human microbiome remains uncertain.Interpretation: The human fecal and respiratory tract microbiome changed in COVID-19 patients with opportunistic pathogen abundance. Further research to elucidate the effect of alternation of the human microbiome in disease pathogenesis is warranted.Funding: This study received no funding.Declaration of Interests: The authors have no conflicts of interest to declare.

Detection of SARS-CoV-2 and Other Infectious Agents in Lower Respiratory Tract Samples Belonging to Patients Admitted to Intensive Care Units of a Tertiary-Care Hospital, Located in an Epidemic Area, during the Italian Lockdown.

The aim of this study was the detection of infectious agents from lower respiratory tract (LRT) samples in order to describe their distribution in patients with severe acute respiratory failure and hospitalized in intensive care units (ICU) in an Italian tertiary-care hospital. LRT samples from 154 patients admitted to ICU from 27 February to 10 May 2020 were prospectively examined for respiratory viruses, including SARS-CoV-2, bacteria and/or fungi. SARS-CoV-2 was revealed in 90 patients (58.4%, 72 males, mean age 65 years). No significant difference was observed between SARS-CoV-2 positives and SARS-CoV-2 negatives with regard to sex, age and bacterial and/or fungal infections. Nonetheless, fungi were more frequently detected among SARS-CoV-2 positives (44/54, 81.4%, p = 0.0053). Candida albicans was the overall most frequently isolated agent, followed by Enterococcus faecalis among SARS-CoV-2 positives and Staphylococcus aureus among SARS-CoV-2 negatives. Overall mortality rate was 40.4%, accounting for 53 deaths: 37 among SARS-CoV-2 positives (mean age 69 years) and 16 among SARS-CoV-2 negatives (mean age 63 years). This study highlights the different patterns of infectious agents between the two patient categories: fungi were prevalently involved among SARS-CoV-2-positive patients and bacteria among the SARS-CoV-2-negative patients. The different therapies and the length of the ICU stay could have influenced these different patterns of infectious agents.

A New SARS-CoV-2 Variant Poorly Detected by RT-PCR on Nasopharyngeal Samples, with High Lethality

Background: In early January 2021, an outbreak of nosocomial cases of COVID-19 emerged in Western France, with RT-PCR tests repeatedly negative on nasopharyngeal samples but positive on lower respiratory tract samples. Whole genome sequencing (WGS) revealed a new variant, currently defining a novel SARS-CoV-2 lineage: B.1.616. In March, WHO classified this variant as ‘under investigation’ (VUI). We analyzed the characteristics and outcomes of COVID-19 cases related to this new variant.Methods: Clinical, virological, and radiological data were retrospectively collected from medical charts in the two hospitals involved. We enrolled patients with at least one of the following: i) positive SARS-CoV-2 RT-PCR on a respiratory sample; ii) seroconversion with anti-SARS-CoV-2 IgG/IgM; iii) suggestive symptoms and typical features of COVID-19 on chest CT scan. Cases were categorized as either: i) B.1.616; ii) variant of concern (VOC); iii) unknown.Findings: From January 1st to March 24th, 2021, 114 patients fulfilled the inclusion criteria: B.1.616 (n=34), VOC (n=32), and unknown (n=48). B.1.616-related cases were older than VOC-related cases (81 years [73-88], vs 73 years [67-82], P Interpretation: We report a nosocomial outbreak of COVID-19 cases related to a new variant, B.1.616, poorly detected by RT-PCR on nasopharyngeal samples, with high lethality.Funding Statement: None. SW lab is funded by Institut Pasteur, CNRS, Université de Paris, Santé publique France, Labex IBEID (ANR-10-LABX- 62-IBEID), REACTing (Research & Action Emerging Infectious Diseases), and by the H2020 project 101003589 (RECOVER).Declaration of Interests: SW has patents issued and pending for SARS-associated coronavirus diagnostics. All other authors declare no competing interestsEthics Approval Statement: The French infectious diseases society ethics committee (Comité d’Éthique de la Recherche en Maladies Infectieuses et Tropicales, CER-MIT) approved the study (N° COVID 2021-06). Written informed consent was waived.

Different Respiratory Samples for COVID-19 Detection by Standard and Direct Quantitative RT-PCR: A Literature Review.

The most common diagnostic method for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is real-time quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR). Upper respiratory tract samples, including nasopharyngeal swab (NPS), oropharyngeal swab (OPS), saliva and lower respiratory tract samples such as sputum, are the most widely used specimens for diagnosis of SARS-CoV-2 using RT-qPCR. This study aimed to compare the diagnostic performance of different samples for Coronavirus disease 2019 (COVID-19) detection. It was found that NPS, the reference respiratory specimen for COVID-19 detection, is more sensitive than OPS. However, the application of NPS has many drawbacks, including challenging sampling process and increased risk of transmission to healthcare workers (HCWs). Saliva samples can be collected less invasively and quickly by HCWs with less contact or by own patients, and they can be considered as an alternative to NPS for COVID-19 detection by RT-qPCR. Additionally, sputum, which demonstrates higher viral load can be applied in patients with productive coughs and negative results from NPS. Commonly, after viral RNA purification from patient samples, which is time-consuming and costly, RT-qPCR is performed to diagnose SARS-CoV-2. Herein, different approaches including physical (heat inactivation) and chemical (proteinase K treatment) methods, used in RNA extraction free- direct RT-qPCR, were reviewed. The results of direct RT-qPCR assays were comparable to the results of standard RT-qPCR, while cost and time were saved. However, optimal protocol to decrease cost and processing time, proper transport medium and detection kit should be determined.

UK B.1.1.7 (Alpha) variant exhibits increased respiratory replication and shedding in nonhuman primates

ABSTRACT The continuing emergence of SARS-CoV-2 variants calls for regular assessment to identify differences in viral replication, shedding and associated disease. In this study, we compared African green monkeys infected intranasally with either the UK B.1.1.7 (Alpha) variant or its contemporary D614G progenitor. Both variants caused mild respiratory disease with no significant differences in clinical presentation. Significantly higher levels of viral RNA and infectious virus were found in upper and lower respiratory tract samples and tissues from B.1.1.7 infected animals. Interestingly, D614G infected animals showed significantly higher levels of viral RNA and infectious virus in rectal swabs and gastrointestinal tissues. Our results indicate that B.1.1.7 infection in African green monkeys is associated with increased respiratory replication and shedding but no disease enhancement similar to human B.1.1.7 cases.

Prevalence and Antimicrobial Susceptibility Pattern of Bacterial Agents Involved in Lower Respiratory Tract Infection at a Tertiary Care Hospital, Jaipur, Rajasthan, India

Introduction: Lower Respiratory Tract Infection (LRTI) is one of the commonest health problems which is not a single disease but a group of specific infections with varying aetiology and symptomatology. These infections are the leading cause of illness and mortality in children and adults across the world because of different epidemiology, pathogenesis, clinical presentation, and outcomes. Aim: To find out the prevalence, bacteriological profile and antimicrobial susceptibility pattern of clinical isolates causing LRTI. Materials and Methods: This was a laboratory based observational study which was carried out between June 2019 to May 2020. All the lower respiratory tract samples (sputum, Endotracheal (ET) aspirate, Bronchoalveolar Lavage (BAL) etc.,) were collected and sample processing was done for isolation and identification by standard methods and antimicrobial susceptibility testing was done using Kirby Bauer disc diffusion method. The data was analysed in Microsoft Office excel worksheet. Results: A total number of 1364 samples were processed in which 615 were culture positive. ET aspirates 65.04% (n=400) showed higher positivity rate than sputum 32.03% (n=197). In this study, gram positive and gram negative bacteria (GNB) were 4.23% (n=26) and 92.19% (n=567) respectively with yeast recovered in only 3.58% (n=22) of the specimens. Among the isolates, Klebsiella pneumoniae 171 (30.15%) was the most common organism. The overall susceptibility of GNB was highest towards tigecycline (51.85%) followed by gentamicin (38.62%). Conclusion: Specific antibiotic utilisation strategies like antibiotic restriction, combination therapy may help to decrease or prevent the emergence of resistance. There is a need for further community based studies to identify the best treatment protocol for individual patients.

Molecular diagnostic methods for pneumonia: how can they be applied in practice?

Pneumonia represents a major burden in clinical practice. A rapid etiological diagnosis is critical for optimizing the antibiotic use. Owing to the variety of possible pathogens and the time needed for bacterial cultures or usual polymerase chain reaction (PCR) assays, timely and precise diagnosis is a huge challenge. Several new rapid multiplex assays have been developed in the last decade to resolve these issues. This review aims to provide an overview of recent evidence on improvements and limitations of new rapid molecular assays for pneumonia. Several rapid multiplex-PCR assays are commercially available for upper or lower respiratory tract samples, allowing detection of a wide range of respiratory viruses, bacteria, and, in some cases, of several antibiotic resistance genes. Clinical evaluations demonstrated their good correlation with gold-standard assays but their lack of exhaustiveness, especially for hospital-acquired pneumonia. Studies that evaluated their potential benefits on antibiotic use suffered from important weaknesses with conflicting and limited results. New molecular assays may enable improvements in patient management and antibiotic use. Available studies highlight several benefits and the strong interrelations needed between microbiologists and physicians for their implementation and interpretation according to the clinical and epidemiological context.

1486. Phylogenetic and alpha toxin variant analyses of Staphylococcus aureus strains isolated from patients during the SAATELLITE study

BackgroundSuvratoxumab is a human monoclonal antibody that neutralizes S. aureus (SA) alpha toxin (AT). SAATELLITE, a phase 2 study of the safety and efficacy of suvratoxumab for reducing the incidence of SA pneumonia (NCT02296320), was conducted within the consortium for Combatting Bacterial Resistance in Europe.MethodsA total of 304 SA isolates (baseline, onset and last available isolates from suspected serious bacterial infections, SSBIs) collected from the lower respiratory tract samples from 165 subjects during SAATELLITE were subjected to whole genome sequencing.AT gene (hla) sequences were translated and amino acid variation was identified in comparison to the reference SA USA300 FPR3757. Phylogenetic analysis, genomic annotation and ST analysis were performed.AT expression in SA culture supernatants was performed by ELISA. Representative isolates with novel AT subtypes that had not been identified in previous studies were tested for hemolytic activity and suvratoxumab neutralizing activity.Wilcoxon rank sum test and Fisher’s exact test were performed, respectively: a) to compare difference in baseline AT expression in relation to SA pneumonia incidence; b) to evaluate the association between occurrence of AT stop codons and incidence of SA pneumonia at baseline, as well as the association between occurrence of AT stop codons and treatment arms at post baseline.ResultsWe identified a total of 44 sequence types (STs) and 21 unique AT subtypes, 7 of which have not been described previously. No substitutions were located in the suvratoxumab binding region and all novel AT subtypes displaying lytic activity were neutralized by suvratoxumab.We detected stop codons Q113B and W205B in AT sequences in 53 and 2 SA isolates, respectively. We uncovered no significant associations of: 1) baseline AT expression with SA pneumonia incidence [p=0.967]; 2) occurrence of AT gene stop codon with either SA pneumonia incidence [p >0.999] or suvratoxumab treatment [p=0.103; lower frequency of stop codons in suvratoxumab arm versus placebo].ConclusionOur data indicated that: 1) suvratoxumab target region in (AT) remains conserved; 2) suvratoxumab is active against all AT variants identified to date; 3) suvratoxumab did not exert pressure on SA clinical isolates for selection of escape mutants.DisclosuresDavid E. Tabor, PhD, AstraZeneca (Employee, Shareholder) Andrey Tovchigrechko, PhD, AstraZeneca (Employee, Shareholder)KitePharma, a Gilead company (Employee, Shareholder) Bret R. Sellman, PhD, AstraZeneca (Employee, Shareholder) Michael McCarthy, n/a, AstraZeneca (Employee) Kathryn Shoemaker, MS, AstraZeneca (Employee) Hasan S. Jafri, MD, FAAP, AstraZeneca (Employee) Mark T. Esser, PhD, AstraZeneca (Employee) Alexey Ruzin, PhD, AstraZeneca (Employee, Shareholder)

Core warming of coronavirus disease 2019 (COVID-19) patients undergoing mechanical ventilation-A protocol for a randomized controlled pilot study.

Coronavirus disease 2019 (COVID-19), caused by the virus SARS-CoV-2, is spreading rapidly across the globe, with little proven effective therapy. Fever is seen in most cases of COVID-19, at least at the initial stages of illness. Although fever is typically treated (with antipyretics or directly with ice or other mechanical means), increasing data suggest that fever is a protective adaptive response that facilitates recovery from infectious illness. To describe a randomized controlled pilot study of core warming patients with COVID-19 undergoing mechanical ventilation. This prospective single-site randomized controlled pilot study will enroll 20 patients undergoing mechanical ventilation for respiratory failure due to COVID-19. Patients will be randomized 1:1 to standard-of-care or to receive core warming via an esophageal heat exchanger commonly utilized in critical care and surgical patients. The primary outcome is patient viral load measured by lower respiratory tract sample. Secondary outcomes include severity of acute respiratory distress syndrome (as measured by PaO2/FiO2 ratio) 24, 48, and 72 hours after initiation of treatment, hospital and intensive care unit length of stay, duration of mechanical ventilation, and 30-day mortality. Resulting data will provide effect size estimates to guide a definitive multi-center randomized clinical trial. ClinicalTrials.gov registration number: NCT04426344. With growing data to support clinical benefits of elevated temperature in infectious illness, this study will provide data to guide further understanding of the role of active temperature management in COVID-19 treatment and provide effect size estimates to power larger studies.

COVID-19 Tanısında Laboratuvar Testleri

The gold standard for routine microbiological diagnosis of coronavirus disease 2019 (COVID-19) is quantitation of viral RNA in respiratory specimens by reverse-transcription polymerase chain reaction (RT-PCR). Detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) specific IgM and IgG antibodies in patient sera are additional diagnostic tests. It has been known that virus release begins a few days before clinical signs appear, and therefore, beginning from 2-3 days before the manifestation of clinical symptoms, virus RNA can be detected in the respiratory tract during the symptomatic period of the disease. Since the viral load is higher in lower respiratory tract samples such as bronchoalveolar lavage and tracheal aspirate, PCR positivity rate might be found higher compared to nasopharyngeal samples. Confirmatory PCR tests require specific equipment and trained personnel, and they are also time-consuming and costly. Antibody assays are simple, faster tests, do not require much equipment and applicable in any laboratory. They can even be performed with 2-3 drops of blood collected from the finger tip of patients using relatively inexpensive chromatographic-rapid tests. These tests can be used in the later period of the disease since specific antibodies appear on the 7-10th day of clinical signs in patients with COVID-19. Rapid antibody card tests have an average specificity and sensitivity, while antibody tests using microELISA have higher sensitivity and specificity.

Pneumocystis jirovecii pneumonia PCR test on upper respiratory tract swab.

Pneumocystis jirovecii pneumonia (PCP) is an opportunistic fungal infection with high morbidity and mortality among people living with HIV. Upper respiratory tract (URT) swabs are routinely taken for testing viral and bacterial pathogens when patients present with respiratory symptoms in our hospital. We conducted a pilot service improvement project to explore the utility of URT swabs for PCP diagnosis using in-house real-time polymerase chain reaction (PCR). Ten URT swab samples obtained from HIV-positive patients with PCP and a positive PCP PCR (AusDiagnostics) from lower respiratory tract (LRT) samples were retrospectively identified. Nine HIV-positive patients with a negative PCR for PCP from LRT samples were identified. Stored aliquots of DNA extracted from these samples were retrieved and tested by an in-house real-time PCR for the presence of PCP DNA. Among PCP-positive cases, URT swabs collected after PCP treatment initiation were excluded from the study. In all, 10 URT samples from PCP-positive patients and nine URT samples from PCP-negative patients were tested for PCP by real-time PCR. Eighteen out of 19 URT sample had a concordant result with the LRT samples. The sensitivity and specificity for URT sample PCR were 90% [confidence interval (CI): 55.50-99.75%] and 100% (CI: 66.37-100%). The positive predictive value was 100% and the negative predictive value was 90.9% (CI: 60.90-98.47%). Upper respiratory tract swab can reliably detect PCP DNA on real-time PCR among people living with HIV with PCP.

Development of a protocol for detection of SARS-CoV-2 in sputum and endotracheal aspirates using Cepheid Xpert Xpress SARS-CoV-2.

Sputum and endotracheal aspirates (ETs) are not among the vendor-approved specimens for the Cepheid Xpert SARS-CoV-2 assay. However, they are the common lower respiratory tract specimens submitted for laboratory diagnosis. Testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in lower respiratory tract samples is required for the discharge of patients from coronavirus disease (COVID) units at some institutions. We developed a protocol used for testing unliquified viscous sputum or tracheal aspirate with the Cepheid Xpert SARS-CoV-2 assay.