Laboratory Diagnosis of COVID-19: Current Issues and Challenges.

The COVID-19 outbreak has had a major impact on clinical microbiology laboratories in the past several months. This commentary covers current issues and challenges for the laboratory diagnosis of infections caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In the preanalytical stage, collecting the proper respiratory tract specimen at the right time from the right anatomic site is essential for a prompt and accurate molecular diagnosis of COVID-19. Appropriate measures are required to keep laboratory staff safe while producing reliable test results. In the analytic stage, real-time reverse transcription-PCR (RT-PCR) assays remain the molecular test of choice for the etiologic diagnosis of SARS-CoV-2 infection while antibody-based techniques are being introduced as supplemental tools. In the postanalytical stage, testing results should be carefully interpreted using both molecular and serological findings. Finally, random-access, integrated devices available at the point of care with scalable capacities will facilitate the rapid and accurate diagnosis and monitoring of SARS-CoV-2 infections and greatly assist in the control of this outbreak.

Comparison of influenza viral load in nasopharyngeal and midturbinate swabs.

Different specimen types are used for influenza diagnosis but comparative data for viral loads from different swab types are limited. We compared influenza viral loads (determined by quantitative reverse transcription polymerase chain reaction [RT-PCR]) in 93 paired midturbinate and nasopharyngeal swab aliquots from influenza infected patients enrolled in a phase 3 randomized-controlled study with the objective of maximizing the number of swabs available for sequence analysis. Midturbinate swabs yielded a 53% lower viral load versus nasopharyngeal swabs, and this difference was similar for influenza A and B. These data suggest that nasopharyngeal swabs might be preferred in diagnostic settings when obtaining higher viral load is important.

Preanalytical Challenges of Molecular Microbiology Tests.

As infectious disease diagnostics increasingly incorporates molecular techniques, there are unique preanalytical concerns that must be considered. First, noninvasive specimen types that may be inadequate for culture-based diagnostics may be acceptable when using molecular tests. Second, specimen containers must be evaluated for the presence of substances that may interfere with amplification or sequencing reactions. Finally, the capacity of transport, storage, and processing conditions to maintain nucleic acid integrity and avoid contamination must be assessed. This review explores these issues and the effects they may have on result quality.

Retrospective Analysis of Sensitivity Characteristics of Enterobacteriaceae: A Study Based on Specimen Types, Sex, and Age Bracket of Patients.

Objective: Enterobacteriaceae have displayed widespread trends of antimicrobial resistance in recent years. Therefore, we aimed to analyse the antimicrobial susceptibility of common bacteria and explore the significance in treatment and research of infections induced by Enterobacteriaceae. Methods: We retrospectively analysed 10,775 antimicrobial susceptibility test results acquired over a 6-year period in the affiliated hospital of Chengde Medical University. We divided the data based on specimen type (blood, sputum, pus, or urine), and population characteristics (age bracket and sex) for analysis. We mainly analysed the antimicrobial susceptibility of Escherichia coli (Eco), Klebsiella pneumoniae (Kpn), and Enterobacter cloacae (Ecl). Results: In our study, it was found that the resistance rates of Eco, Kpn, and Ecl to most antimicrobial agents were significantly different (P < 0.05) on specimen type and age bracket. The Eco from sputum had the highest resistance rates except ciprofloxacin (CIP), levofloxacin (LVX), and gentamicin (GEN); the Kpn from urine had the highest resistance rates to all antimicrobial agents; the Ecl from urine had the highest resistance rates to most antimicrobial agents. The Eco from geriatric patients had the highest resistance rates except GEN and SXT; the Kpn from adult patients had the lowest resistance rates to most antimicrobial agents except LVX. The Eco isolated from males had higher resistance rates to most antimicrobial agents except CIP, LVX, and NIT than those isolated from females; the Kpn showed significant differences in antimicrobial susceptibility to only 5 out of 22 antimicrobial agents (P < 0.05); the Ecl showed significant differences in susceptibility only to two antimicrobial agents, LVX and TOB (P < 0.01). Conclusion: The antimicrobial susceptibility of Enterobacteriaceae was significantly different among specimen type, age bracket and sex of patients, which is of great significance for the treatment and research of infection.

Rapid diagnostic testing for syphilis in Arctic communities (the STAR study): a multisite prospective field diagnostic accuracy study in an intended-use setting.

OBJECTIVES: We evaluated the field diagnostic accuracy of a syphilis rapid test (RDT), using serum and whole blood by non-laboratorians in two Canadian Arctic communities. METHODS: We implemented a multisite prospective field evaluation wherein patients were screened by an RDT containing treponemal and non-treponemal components (Chembio DPP® Syphilis Screen & Confirm) between January 2020 and December 2021. Venous whole blood and serum were collected for rapid testing and compared with laboratory-based serology reference testing using a reverse sequence algorithm of treponemal and rapid plasma reagin (RPR) testing. RESULTS: Overall, 135 whole blood and 139 serum specimens were collected from 161 participants during clinical encounters. Treponemal-RDT sensitivity against a treponemal-reference standard (38/161 confirmed cases) was similar for serum (78% [95% CI: 61-90%]) and whole blood (81% [95% CI: 63-93%]). In those with RPR titres ≥1:8 (i.e. suggestive of recent/active infection), sensitivity increased to 93% (95% CI: 77-99%) for serum and 92% (95% CI: 73-99%) for whole blood. Treponemal-RDT specificity was excellent (99% [95% CI: 95-100%]) for both specimen types. Non-treponemal-RDT sensitivity against RPR was 94% (95% CI: 80-99%) for serum and 79% (95% CI: 60-92%) for whole blood. Sensitivity increased to 100% (95% CI: 88-100%) for serum and 92% (95% CI: 73-99%) for whole blood when RPR titres ≥1:8. RDT performance with whole blood was similar to that with serum. DISCUSSION: Non-laboratorians using the RDT accurately identified individuals with infectious syphilis under real-world conditions in an intended-use setting at the point of care. Implementing the RDT can eliminate treatment delays and may enhance disease control.

Better choice of the type of specimen used for untargeted metagenomic sequencing in the diagnosis of periprosthetic joint infections.

AIMS: As a proven and comprehensive molecular technique, metagenomic next-generation sequencing (mNGS) has shown its potential in the diagnosis of pathogens in patients with periprosthetic joint infection (PJI), using a single type of specimen. However, the optimal use of mNGS in the management of PJI has not been explored. In this study, we evaluated the diagnostic value of mNGS using three types of specimen with the aim of achieving a better choice of specimen for mNGS in these patients. METHODS: In this prospective study, 177 specimens were collected from 59 revision arthroplasties, including periprosthetic tissues, synovial fluid, and prosthetic sonicate fluid. Each specimen was divided into two, one for mNGS and one for culture. The criteria of the Musculoskeletal Infection Society were used to define PJI (40 cases) and aseptic failure (19 cases). RESULTS: The sensitivity and specificity of mNGS in the diagnosis of PJI were 95% and 94.7%, respectively, for all types of specimen. The sensitivity and specificity were 65% and 100%, respectively, for periprosthetic tissues, 87.5% and 94.7%, respectively, for synovial fluid, and 92.5% and 94.7%, respectively, for prosthetic sonicate fluid. The mNGS of prosthetic sonicate fluid outperformed that for other types of specimen in the rates of detection of pathogens (84.6%), sequencing reads (> ten-fold) and the rate of genome coverage (> five-fold). CONCLUSION: mNGS could serve as an accurate diagnostic tool in the detection of pathogens in patients with a PJI using three types of specimen. Due to its superior perfomance in identifying a pathogen, mNGS of prosthetic sonicate fluid provides the most value and may partly replace traditional tests such as bacteriological culture in these patients. Cite this article: Bone Joint J 2021;103-B(5):923-930.

Analysis of NTRK mutation and clinicopathologic factors in lung cancer patients in northeast China.

OBJECTIVE: NTRK mutations and clinicopathological factors in patients with lung cancer in northeast China were analyzed by next-generation sequencing (NGS), and references were provided for patients with NTRK mutations undergoing targeted therapy in northeast China. METHODS: A total of 224 specimens in 173 patients with lung cancer were collected. This included 51 patients with matched tissue and whole blood samples,133 tissue samples, 84 whole blood samples, and 7 pleural effusion samples. NGS (520 genes) was used to detected NTRK mutations and clinicopathologic factors. RESULTS: NTRK mutation was detected in eight patients (8/173, 4.6%), including four NTRK missense mutations (4/173, 2.3%), two NTRK fusion gene mutations (2/173, 1.2%), and two NTRK copy number deletions (2/173, 1.2%). Among the eight patients with NTRK mutations, four were associated with lung cancer driver gene mutations (3/4 EGFR, 1/4ALK); NTRK in two patients was inconsistent in tissue and paired whole blood testing; NTRK missense mutation was detected in one patient, and NTRK copy number deletion was detected in the other; and NTRK wild type was detected in two patients. There was no correlation between NTRK mutation and clinicopathologic factors (including gender, age, pathological type, smoking status, metastasis site). CONCLUSION: NTRK mutation was only 4.6%, effective fusion gene mutation was 1.2%, and common driver gene mutation in lung cancer was evident in 50% of patients. The results of NTRK were inconsistent with matched tissues and whole blood. Therefore, patients with NTRK mutation should use a variety of specimen types and large target area sequencing (panel) analysis method to provide individualized treatment.

Validation of SARS-CoV-2 detection across multiple specimen types.

BACKGROUND: Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has caused considerable disruption across the world, resulting in more than 235,000 deaths since December 2019. SARS-CoV-2 has a wide tropism and detection of the virus has been described in multiple specimen types, including various respiratory secretions, cerebrospinal fluid, and stool. OBJECTIVE: To evaluate the accuracy and sensitivity of a laboratory modified CDCbased SARS-CoV-2 N1 and N2 assay across a range of sample types. Study Design We compared the matrix effect on the analytical sensitivity of SARS-CoV-2 detection by qRT-PCR in nasal swabs collected in viral transport medium (VTM), bronchoalveolar lavage (BAL), sputum, plasma, cerebral spinal fluid (CSF), stool, VTM, phosphate buffered saline (PBS), and Hanks' Balanced Salt Solution (HBSS). Initial limits of detection (LoD) were subsequently narrowed to confirm an LoD for each specimen type and target gene. RESULTS: LoDs were established using a modified CDC-based laboratory developed test and ranged from a mean CT cut-off of 33.8-35.7 (10-20 copies/reaction) for the N1 gene target, and 34.0-36.2 (1-10 copies/reaction) for N2. Alternatives to VTM such as PBS and HBSS had comparable LoDs. The N2 gene target was found to be most sensitive in CSF. CONCLUSION: A modified CDC-based laboratory developed test is able to detect SARSCoV- 2 accurately with similar sensitivity across all sample types tested.

Three new synonyms within the flower chafer genus Goliathopsis Janson, 1881 (Coleoptera: Scarabaeidae: Cetoniinae) from China.

All available type materials of the Indochinese genus Goliathopsis Janson, 1881 are determined, and lectotype designations are made for G. capreolus Gestro 1888, G. despectus (Westwood, 1873), G. esquiroli Pouillaude, 1913, and G. velutinus Pouillaude, 1913. The examination of type series revealed three synonyms: Goliathopsis camptotropus Yang, 1988 new synonym and G. polystricus Yang, 1988 new synonym are synonymized with G. esquiroli, and G. maolanus Yang, 1988 new synonym is regarded as a junior synonym of G. lameyi Fairmaire, 1893. Besides, G. cervus Janson, 1881 is proved to be a valid species. Diagnoses, key, and color illustrations are provided for all species. All known distribution records are mapped, and the location of "Tsékou" and "Oui-Sy" are presented and discussed.

World Catalogue of the Druid Flies (Diptera: Schizophora: Clusiidae).

Family-, genus- and species-level groups in the family Clusiidae (Diptera: Schizophora) are catalogued, providing reference to occurrences of these taxa in the literature, including all those relevant to nomenclature. Full synonymies are provided, including generic combinations for species, and the collection locality, depository and sex of primary type specimens. Published species distributions are provided, noting country and biogeographic region; specimen data representing new country records for species are listed. The Clusiidae are known from 636 species in 14 genera and three subfamilies, with many additional species expected. Nearly half of all described species are Sobarocephala, with 269 species, followed by the clusiodine genera Heteromeringia (86 species), Allometopon (68), Hendelia (53 species), Czernyola (50 species) and Clusiodes (31 species); the remaining eight genera (seven extant, one fossil) are smaller, consisting of 1 to 16 species. Two other fossil genera, Acartophthalmites Hennig and Xenanthomyza Hennig, are also treated here, but are unlikely to belong to the family. Taxa formerly considered Clusiidae are listed. Tranomeringia scutellata Sasakawa is transferred to Heteromeringia, n. comb. Czernyola is maintained as the replacement name for Craspedochaeta, following Bezzi (1907), McAlpine (1971) and Lonsdale, et al. (2010), resulting in the following new combinations: Czernyola amazonensis (Lonsdale & Marshall) n. comb.; C. apsilutea (Lonsdale & Marshall) n. comb.; C. argoniae (Lonsdale & Marshall) n. comb.; C. biloba (Lonsdale & Marshall) n. comb.; C. brunneivibrissa (Lonsdale & Marshall) n. comb.; C. candida (Lonsdale & Marshall) n. comb.; C. chauliodon (Lonsdale & Marshall) n. comb.; C. chela (Lonsdale & Marshall) n. comb.; C. concinna (Williston) n. comb.; C. feminea (Lonsdale & Marshall) n. comb.; C. loreto (Lonsdale & Marshall) n. comb.; C. maai (Sasakawa) n. comb.; C. melanosoma (Lonsdale & Marshall) n. comb.; C. novaeguinea (Soós) n. comb.; C. pacaraima (Lonsdale & Marshall) n. comb.; C. parva (Sasakawa) n. comb.; C. phaios (Lonsdale & Marshall) n. comb.; C. pilosa (Sasakawa) n. comb.; C. pleuralis (Williston) n. comb.; C. pollostos (Lonsdale & Marshall) n. comb.; C. protomis (Lonsdale & Marshall) n. comb.; C. quinquespinula (Sasakawa) n. comb.; C. sasakawai (Lonsdale & Marshall) n. comb.; C. spinulifera (Sasakawa) n. comb.; C. unguicauda (Sasakawa) n. comb.; C. varicolor (Sueyoshi) n. comb.; C. vietnamensis (Sasakawa) n. comb.; C. weemsi (Lonsdale & Marshall) n. comb.; C. xanthonotum (Lonsdale & Marshall) n. comb.; C. xanthopleura (Sasakawa) n. comb.; C. zongo (Lonsdale & Marshall) n. comb.

Epstein-Barr virus serology and PCR: conflicting results in an immunocompetent host. A case report and review of literature.

We present the case of a 27-year-old immunocompetent man who progressively developed a generalized lymphadenopathy and B symptoms. Results of Epstein-Barr virus (EBV) serology were suggestive for a past infection, but the EBV viral load in whole blood was high. Also, core needle biopsy of the largest lymph node showed an image which could fit an EBV-driven reactive lymphoproliferation. Despite the absence of an immune disorder, all medical evidence points to an EBV-driven lymphoproliferative proces. In immunocompetent patients, it seems extremely uncommon to detect a high EBV viral load in the absence of serological evidence of an acute EBV infection or reactivation. We reviewed literature on this topic and on the selection of the appropriate sample type for EBV PCR, as this is known to be a critical point. Serological testing for the diagnosis of EBV infection is the gold standard in immunocompetent patients. Measuring EBV viral load is only recommended when dealing with immunocompromised patients. Although extremely rare, this case report shows that there is a place for EBV PCR in certain situations in immunocompetent patients. Besides, there is still no consensus regarding the specimen of choice for the determination of the EBV viral load. The preferred specimen type seems to depend on the patient's underlying condition.