COVID-19 surveillance in a bone marrow transplantation unit: experience from a Brazilian tertiary-care teaching hospital.

PURPOSE: In this work, we aimed to describe the strategy of the weekly SARS-CoV-2 RT-PCR surveillance program that was implemented in our bone marrow transplantation (BMT) unit. METHODS: Our unit performed SARS-CoV-2 RT-PCR before admission and then weekly during hospitalization even if the patient was asymptomatic. From May 2021 to May 2022, we collected data from all patients that were admitted in the BMT unit to perform transplantation. The total of SARS-CoV-2 RT-PCR performed and the positive rate were described. RESULTS: During the study period, 65 patients were admitted for HSCT. A total of 414 SARS-CoV-2 RT-PCR were performed. Two cases were detected (positivity rate, 0.48%). After the positive test, both patients were isolated outside the BMT unit. CONCLUSION: We postulate that diagnosing these patients and isolating them outside the transplantation unit may have prevented secondary symptomatic cases.

Clinical and laboratory diagnosis of Mayaro virus (MAYV): Current status and opportunities for further development.

The recent outbreaks related to Mayaro virus (MAYV) infection in the Americas have brought this neglected virus as a potential threat to global public health. Given the range of symptoms that can be associated with MAYV infection, it can be challenging to diagnose individuals based on clinical signs, especially in countries with simultaneous circulation of other mosquito-borne viruses, such as dengue virus (DENV) and chikungunya virus (CHIKV). With this challenge in mind, laboratory-based diagnosis assumes a critical role in the introduction of measures to help prevent virus dissemination and to adequately treat patients. In this review, we provide an overview of the clinical features reported in infected patients and currently available laboratory tools that are used for MAYV diagnosis, discussing their advances, advantages, and limitations to apply in the field. Moreover, we explore novel point-of-care (PoC) diagnostic platforms that can provide de-centralised diagnostics for use in areas with limited laboratory infrastructure.

Improved heat shock method for extracting total RNA from nasopharyngeal swab samples even with low viral load.

This study aimed to improve the heat shock method as a cost-effective and time-efficient for total RNA extraction. We compared the effectiveness of two total RNA extraction methods by using Real-Time PCR for nasopharynx swabs. Include: I; use of a commercial total RNA extraction kit as a standard. II; utilized a modified heat shock method (MHS). Time, centrifuge speed and duration, proteinase K, and RNA carrier were optimized. The optimized parameters included treating the sample with 5 µg/µL at 56°C for 5 minutes, heating at 95°C for 5 minutes followed by thermal shock in ice for 3 minutes, adding 4 µg/µL RNA carrier at room temperature for 3 minutes, and centrifuging at 7000 rpm for 10 minutes. This optimization demonstrated a sensitivity and specificity of 100% (CI: 95%) even in samples with low viral load. Our in-house method presents a rapid, and cost-effective alternative for total RNA extraction.

Evaluation of a rapid Loop Mediated Isothermal Amplification (LAMP) test for the laboratory diagnosis of sexually transmitted infections.

Sexually transmitted infections (STIs) have seen a considerable increase in the last years and given the health burden they may represent from both a personal and community perspective, they require surveillance and prevention programmes based on a timely and decentralized diagnosis. In this context, user-friendly rapid molecular tests may represent a good trade-off between diagnostic accuracy, accessibility and affordability. In this study we evaluated the diagnostic performance of a new real-time LAMP (Loop Mediated Isothermal Amplification) method for the rapid detection and differentiation of 7 major sexually transmissible pathogens by analysing real clinical samples (genital and extra-genital matrices) from individuals with suspected STIs. The assay showed good overall diagnostic performances in terms of sensitivity, specificity and concordance with a gold-standard PCR-based molecular method. This assay, not requiring specialised laboratory technicians or expensive instrumentation, but nonetheless capable of guaranteeing accurate results, is within the reach of outpatient settings, obstetrics, and gynaecology clinic, hence ensuring on-field access to early diagnosis.

Clinical validation of RCSMS: A rapid and sensitive CRISPR-Cas12a test for the molecular detection of SARS-CoV-2 from saliva.

Peru's holds the highest COVID death rate per capita worldwide. Key to this outcome is the lack of robust, rapid, and accurate molecular tests to circumvent the elevated costs and logistics of SARS-CoV-2 detection via RT-qPCR. To facilitate massive and timely COVID-19 testing in rural and socioeconomically deprived contexts, we implemented and validated RCSMS, a rapid and sensitive CRISPR-Cas12a test for the molecular detection of SARS-CoV-2 from saliva. RCSMS uses the power of CRISPR-Cas technology and lateral flow strips to easily visualize the presence of SARS-CoV-2 even in laboratories with limited equipment. We show that a low-cost thermochemical treatment with TCEP/EDTA is sufficient to inactivate viral particles and cellular nucleases in saliva, eliminating the need to extract viral RNA with commercial kits, as well as the cumbersome nasopharyngeal swab procedure and the requirement of biosafety level 2 laboratories for molecular analyses. Notably, RCSMS performed outstandingly in a clinical validation done with 352 patients from two hospitals in Lima, detecting as low as 50 viral copies per 10 µl reaction in 40 min, with sensitivity and specificity of 96.5% and 99.0%, respectively, relative to RT-qPCR. The negative and positive predicted values obtained from this field validation indicate that RCSMS can be confidently deployed in both high and low prevalence settings. Like other CRISPR-Cas-based biosensors, RCSMS can be easily reprogrammed for the detection of new SARS-CoV-2 variants. We conclude that RCSMS is a fast, efficient and inexpensive alternative to RT-qPCR for expanding COVID-19 testing capacity in Peru and other low- and middle-income countries with precarious healthcare systems.

External quality assurance in the era of standardization.

Metrology in clinical chemistry aims to ensure the equivalence of measurement results from different in-vitro diagnostic measurement devices (IVD MD) for use in healthcare. The metrological traceability of measurement results to higher-order references is the cornerstone to achieving equivalent results. However, other fundamentals are also needed, including the commutability of reference materials and external quality assessment (EQA) materials for monitoring the equivalence of measurement results at the end-user level. This manuscript summarizes the findings and opinions expressed at the Joint Community for Traceability in Laboratory Medicine (JCTLM) workshop held on December 4-5, 2023. The workshop explored the relationship between EQA/proficiency testing and metrological traceability to higher-order references. EQA monitors the equivalence of measurement results from end-user IVD MDs. The workshop discussed the role and challenges of using EQA to improve and maintain the equivalence of measurement results. It also elucidated current developments in establishing the clinical suitability of laboratory results expressed as analytical performance specifications (APS).

A Generalizable Decision-Making Framework for Selecting Onsite versus Send-out Clinical Laboratory Testing.

BACKGROUND: Laboratory networks provide services through onsite testing or through specimen transport to higher-tier laboratories. This decision is based on the interplay of testing characteristics, treatment characteristics, and epidemiological characteristics. OBJECTIVES: Our objective was to develop a generalizable model using the threshold approach to medical decision making to inform test placement decisions. METHODS: We developed a decision model to compare the incremental utility of onsite versus send-out testing for clinical purposes. We then performed Monte Carlo simulations to identify the settings under which each strategy would be preferred. Tuberculosis was modeled as an exemplar. RESULTS: The most important determinants of the decision to test onsite versus send-out were the clinical utility lost due to send-out testing delays and the accuracy decrement with onsite testing. When the sensitivity decrements of onsite testing were minimal, onsite testing tended to be preferred when send-out delays reduced clinical utility by >20%. By contrast, when onsite testing incurred large reductions in sensitivity, onsite testing tended to be preferred when utility lost due to delays was >50%. The relative cost of onsite versus send-out testing affected these thresholds, particularly when testing costs were >10% of treatment costs. CONCLUSIONS: Decision makers can select onsite versus send-out testing in an evidence-based fashion using estimates of the percentage of clinical utility lost due to send-out delays and the relative accuracy of onsite versus send-out testing. This model is designed to be generalizable to a wide variety of use cases. HIGHLIGHTS: The design of laboratory networks, including the decision to place diagnostic instruments at the point-of-care or at higher tiers as accessed through specimen transport, can be informed using the threshold approach to medical decision making.The most important determinants of the decision to test onsite versus send-out were the clinical utility lost due to send-out testing delays and the accuracy decrement with onsite testing.The threshold approach to medical decision making can be used to compare point-of-care testing accuracy decrements with the lost utility of treatment due to send-out testing delays.The relative cost of onsite versus send-out testing affected these thresholds, particularly when testing costs were >10% of treatment costs.

Behavioral Intention of Receiving Monkeypox Vaccination and Undergoing Monkeypox Testing and the Associated Factors Among Young Men Who Have Sex With Men in China: Large Cross-Sectional Study.

BACKGROUND: The worldwide human monkeypox (mpox) outbreak in 2022 mainly affected men who have sex with men (MSM). In China, young men who have sex with men (YMSM) were at a potential high risk of mpox infection due to their sexual activeness and the eased COVID-19 restrictions at the end of 2022. OBJECTIVE: This study aimed to investigate the behavioral intention of receiving mpox vaccination and undergoing mpox testing in 4 different scenarios and explore their associations with background and behavioral theory-related factors among Chinese YMSM. METHODS: An online cross-sectional survey was conducted among YMSM aged 18-29 years from 6 representative provinces of China in September 2022. Participants recruited (recruitment rate=2918/4342, 67.2%) were asked to self-administer an anonymous questionnaire designed based on prior knowledge about mpox and classic health behavior theories. Data on the participants' background, mpox knowledge and cognition, mpox vaccination and testing cognition, and the behavioral intention of receiving mpox vaccination and undergoing mpox testing were collected. Descriptive analysis and univariate and multivariate linear regressions were performed. Geodetector was used to measure the stratified heterogeneity of behavioral intention. RESULTS: A total of 2493 YMSM with a mean age of 24.6 (SD 2.9) years were included. The prevalence of having a behavioral intention of receiving mpox vaccination ranged from 66.2% to 88.4% by scenario, varying in epidemic status and cost. The prevalence of having an mpox testing intention was above 90% in all scenarios regardless of the presence of symptoms and the cost. The positive factors related to vaccination intention included mpox knowledge (ba=0.060, 95% CI 0.016-0.103), perceived susceptibility of mpox (ba=0.091, 95% CI 0.035-0.146), perceived severity of mpox (ba=0.230, 95% CI 0.164-0.296), emotional distress caused by mpox (ba=0.270, 95% CI 0.160-0.380), perceived benefits of mpox vaccination (ba=0.455, 95% CI 0.411-0.498), self-efficacy of mpox vaccination (ba=0.586, 95% CI 0.504-0.668), and having 1 male sex partner (ba=0.452, 95% CI 0.098-0.806), while the negative factor was perceived barriers to vaccination (ba=-0.056, 95% CI -0.090 to -0.022). The positive factors related to testing intention were perceived severity of mpox (ba=0.283, 95% CI 0.241-0.325), perceived benefits of mpox testing (ba=0.679, 95% CI 0.636-0.721), self-efficacy of mpox testing (ba=0.195, 95% CI 0.146-0.245), having 1 male sex partner (ba=0.290, 95% CI 0.070-0.510), and having in-person gatherings with MSM (ba=0.219, 95% CI 0.072-0.366), while the negative factor was emotional distress caused by mpox (ba=-0.069, 95% CI -0.137 to -0.001). CONCLUSIONS: Among Chinese YMSM, the intention of undergoing mpox testing is optimal, while the mpox vaccination intention has room for improvement. A future national response should raise YMSM's mpox knowledge, disseminate updated information about mpox and preventive measures, improve preventive service accessibility and privacy, and provide advice on positively coping with the associated emotional distress.

Comparative evaluation of assay performance for SARS-CoV-2 detection in animal oral samples, lung homogenates, and phosphate-buffered saline using the TaqPath COVID-19 Combo kit.

A One Health approach has been key to monitoring the COVID-19 pandemic, as human and veterinary medical professionals jointly met the demands for an extraordinary testing effort for SARS-CoV-2. Veterinary diagnostic laboratories continue to monitor SARS-CoV-2 infection in animals, furthering the understanding of zoonotic transmission dynamics between humans and animals. A RT-PCR assay is a primary animal screening tool established within validation and verification guidelines provided by the American Association of Veterinary Laboratory Diagnosticians (AAVLD), World Organisation for Animal Health (WOAH), and the U.S. Food and Drug Administration (FDA). However, differences in sample matrices, RNA extraction methods, instrument platforms, gene targets, and cutoff values may affect test outcomes. Therefore, targeted validation for a new sample matrix used in any PCR assay is critical. We evaluated a COVID-19 assay for the detection of SARS-CoV-2 in feline and canine lung homogenates and oral swab samples. We used the commercial Applied Biosystems MagMAX Viral/Pathogen II (MVP II) nucleic acid isolation kit and TaqPath COVID-19 Combo kit, which are validated for a variety of human samples, including nasopharyngeal and oropharyngeal swab samples. Our masked test showed a high detection rate and no false-positive or false-negative results, supporting sample extension to include feline oral swab samples. Our study is a prime example of One Health, illustrating how a COVID-19 assay designed for human testing can be adapted and used to detect SARS-CoV-2 in oral swab samples from cats and likely dogs, but not lung homogenates.

Recent updates on laboratory diagnosis of rabies.

Rabies is a lethal viral disease transmitted through the bite of rabid animals. India has a high burden of rabies, contributing to a significant proportion of the global deaths. However, under-reporting of the disease is prevalent due to lack of laboratory confirmation. Laboratory diagnosis of rabies plays a crucial role in differentiating the disease from clinical mimics, initiation of appropriate care, implementing infection control measures and informing disease surveillance. This review provides an overview of the recent advancements in laboratory diagnosis of rabies, aimed at updating physicians involved in diagnosis and management of rabies cases in India.

Nationwide quality assurance of high-throughput diagnostic molecular testing during the SARS-CoV-2 pandemic: role of the Belgian National Reference Centre.

Since the onset of the coronavirus disease (COVID-19) pandemic in Belgium, UZ/KU Leuven has played a crucial role as the National Reference Centre (NRC) for respiratory pathogens, to be the first Belgian laboratory to develop and implement laboratory developed diagnostic assays for SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) and later to assess the quality of commercial kits. To meet the growing demand for decentralised testing, both clinical laboratories and government-supported high-throughput platforms were gradually deployed across Belgium. Consequently, the role of the NRC transitioned from a specialised testing laboratory to strengthening capacity and coordinating quality assurance. Here, we outline the measures taken by the NRC, the national public health institute Sciensano and the executing clinical laboratories to ensure effective quality management of molecular testing throughout the initial two years of the pandemic (March 2020 to March 2022).

Competency of malaria laboratory diagnosis at national and provincial levels at the beginning of malaria post-elimination phase, China.

BACKGROUND: Qualified malaria diagnosis competency has contributed to the great achievement of malaria elimination in China. After eliminating malaria, it is still critical to the prevention of re-establishment of malaria transmission in China. This study was aimed to assess the malaria detection competency at national and provincial levels in China at the beginning of malaria post-elimination phase. METHODS: In the present study, different competency assessment activities on the laboratory malaria diagnosis were carried out for national and provincial malaria diagnostic laboratories based on the WHO scoring schedules, including malaria microscopy or nucleic acid amplification tests (NAAT), at the beginning of malaria post-elimination phase (2021-2022) in China. RESULTS: A total of 60 slides for malaria microscopy and 10 specimen for NAAT were included into the WHO External Quality Assessments of malaria parasite qualitative detection and species identification, and the scoring rate was 96.6% (microscopy: 171/177) and 85.0% (NAAT: 17/20), respectively. Moreover, 124 samples were included into the national NAAT quality assessment, and an accuracy of 87.9% (109/124) was found without significance among reference laboratories and non-reference laboratories. CONCLUSIONS: The findings suggest that there is still a need for sustained strengthening of malaria detection competency, particularly in the areas of parasite counting and detection of low-density parasitemia, to ensure prompt detection of the sources of infection and accurate identification of Plasmodium species, and contribute to case management and focus disposal, thereby effectively preventing the malaria re-establishment.

Evaluation of the EasyNAT SARS-CoV-2 assay PCR test for the diagnosis of SARS-CoV-2 infection.

Reverse transcription polymerase chain reaction (RT-PCR) tests are commonly utilized in commercial settings but pose challenges due to labor-intensive procedures and extended response times during peak demand. In contrast, real-time fluorescence and isothermal amplification assays using Crossing Priming Amplification (CPA) offer faster genetic material analysis, eliminate subjectivity, and require less manipulation and personnel training. This study aimed to validate the EasyNAT SARS-CoV-2 Assay, a diagnostic kit based on CPA, using oral and nasopharyngeal samples. The EasyNAT kit was compared to the Xpert Xpress SARS-CoV-2 kit, evaluating 873 samples obtained during routine analysis at the Microbiology Laboratory of the Hospital Costa del Sol (Marbella, Spain). The overall sensitivity and specificity for the EasyNAT SARS-CoV-2 Assay were 79.1% (95%CI 74.5-83.7) and 99.5% (95%CI 98.7-100), respectively; with, validity index of 91.9%, positive predictive value of 98.9%, negative predictive value of 88.9%, positive likelihood ratio of 144.5, negative likelihood ratio of 0.21 and a total Youden Index of 0.79. Notably, sensitivity improved in fresh samples (91.4%), along with a high Youden Index (0.91). The EasyNAT SARS-CoV-2 Assay achieved a higher percentage of concordance in positive samples with Xpert Xpress SARS-CoV-2 when analyzing cycle threshold (Ct) intervals below 30 compared to intervals equal or greater than 30, and demons. In conclusion, the EasyNAT SARS-CoV-2 Assay demonstrated high sensitivity and agreement with Xpert Xpress SARS-CoV-2, particularly in fresh samples or when the signal was detected at Ct intervals below 30, indicating higher viral loads. This makes it suitable for rapid screening in various settings, including those with limited access to conventional molecular laboratory setting.

The Laboratory Diagnosis of Malaria: A Focus on the Diagnostic Assays in Non-Endemic Areas.

Even if malaria is rare in Europe, it is a medical emergency and programs for its control should ensure both an early diagnosis and a prompt treatment within 24-48 h from the onset of the symptoms. The increasing number of imported malaria cases as well as the risk of the reintroduction of autochthonous cases encouraged laboratories in non-endemic countries to adopt diagnostic methods/algorithms. Microscopy remains the gold standard, but with limitations. Rapid diagnostic tests have greatly expanded the ability to diagnose malaria for rapid results due to simplicity and low cost, but they lack sensitivity and specificity. PCR-based assays provide more relevant information but need well-trained technicians. As reported in the World Health Organization Global Technical Strategy for Malaria 2016-2030, the development of point-of-care testing is important for the improvement of diagnosis with beneficial consequences for prompt/accurate treatment and for preventing the spread of the disease. Despite their limitations, diagnostic methods contribute to the decline of malaria mortality. Recently, evidence suggested that artificial intelligence could be utilized for assisting pathologists in malaria diagnosis.

The diagnostic accuracy of the ID NOW COVID-19 point of care test in acute hospital admissions.

BACKGROUND: Prompt identification of patients with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection on admission to hospital is crucial to ensuring initiation of appropriate treatment, optimising infection control and maintaining patient flow. The Abbott ID NOW™ COVID-19 assay (ID NOW) is a point-of-care, isothermal nucleic acid amplification test, capable of producing a result within minutes, potentially placing it as an invaluable tool in helping to control the coronavirus-disease 2019 (COVID-19) pandemic. OBJECTIVES: To evaluate the diagnostic accuracy of ID NOW in acute hospital admissions. STUDY DESIGN: A prospective approach to data collection was undertaken in consecutive patients with ID NOW and Hologic Aptima™ SARS-CoV-2 transcription-mediated amplification assay (Aptima TMA) results, across three hospitals in the south-west of England between 1st March and 30th September 2021. A nasal swab was taken for ID NOW and a combined nose and throat swab for Aptima TMA. Measures of diagnostic accuracy were calculated for ID NOW against Aptima TMA. This study was conducted during a period of alpha and delta strain predominance. RESULTS: 19,698 ID NOW assays were performed, of which 12,821 had an Aptima TMA assay performed within 24 hours. ID NOW had sensitivity of 85.2 % (95 % CI, 82.2-87.9) and specificity of 99.6 % (95 % CI, 99.4-99.7) compared with the reference assay. The overall PPV was 91.0 % (95 % CI, 88.5-93.0) and the overall NPV was 99.3 % (95 % CI, 99.1-99.4). CONCLUSIONS: ID NOW offers a valid diagnostic tool to detect SARS-CoV-2, performing comparably to a reference laboratory-based assay which takes longer to provide results.

Home and Clinical Laboratory Improvement Amendments-Waived Testing for Infectious Diseases-How Do These Fit in the Testing Landscape?

Though testing for infectious diseases has long been performed in traditional clincial laboratory settings, more widespread availability of waived testing is expanding accessibility of patients to rapid test results. This is being further expanded to home testing. Nevertheless, with this greater democratization and availability of clinical testing there are important limitations that need to be balanced. In this article, we review the current test landscape for infectious diseases waived testing and opportunities for assuring optimal quality testing.

ICSH review of internal quality control policy for blood cell counters.

INTRODUCTION: This paper is a report of an ICSH review of policies and practices for internal quality control (IQC) policy for haematology cell counters among regulatory bodies, cell counter manufacturers and diagnostic laboratories. It includes a discussion of the study findings and links to separate ICSH guidance for such policies and practices. The application of internal quality control (IQC) methods is an essential pre-requisite for all clinical laboratory testing including the blood count (Full Blood Count, FBC, or Complete Blood Count, CBC). METHODS: The ICSH has gathered information regarding the current state of practice through review of published guidance from regulatory bodies, a questionnaire to six major cell counter manufacturers (Abbott Diagnostics, Beckman Coulter, Horiba Medical Diagnostic Instruments & Systems, Mindray Medical International, Siemens Healthcare Diagnostics and Sysmex Corporation) and a survey issued to 191 diagnostic laboratories in four countries (China, Republic of Ireland, Spain and the United Kingdom) on their IQC practice and approach to use of commercial IQC materials. RESULTS: This has revealed diversity both in guidance and in practice around the world. There is diversity in guidance from regulatory organizations in regard to IQC methods each recommends, clinical levels to use and frequency to run commercial controls, and finally recommended sources of commercial controls. The diversity in practice among clinical laboratories spans the areas of IQC methods used, derivation of target values and action limits used with control materials, and frequency of running commercial controls materials. CONCLUSIONS: These findings and their implications for IQC Practice are discussed in this paper. They are used to inform a separate guidance document, which proposes a harmonized approach to address the issues faced by diagnostic laboratories.

Review of virological methods for laboratory diagnosis and characterization of monkeypox virus (MPXV): lessons learned from the 2022 Mpox outbreak.

Monkeypox virus (MPXV), originally endemic in West Africa (Clade II) and Central Africa (Clade I), has recently emerged worldwide and has reinforced the need for rapid and accurate MPXV diagnostics. This review presents and critically discusses the range of virological methods for laboratory diagnosis and characterization of MPXV as well as related lessons learned and practical experience gained from the 2022 Mpox global outbreak. Real-time PCR is currently considered the diagnostic gold standard and ensures accurate and timely confirmation of suspected Mpox cases based on suspicious skin lesions, and digital PCR improves the precision of MPXV DNA quantification. Whole genome sequencing reveals the diversity within the Clade IIb outbreak and highlights the role of microevolution in the adaptation of the virus to the human host. Continuous genomic surveillance is important for better understanding of human-to-human transmission and prevention of the emergence of variola virus-like strains. Traditional virological methods such as electron microscopy and virus isolation remain essential for comprehensive virus characterization, particularly in the context of vaccine and antiviral drug development. Despite the current challenges, serological tests detecting a range of anti-MPXV antibodies are important adjunct diagnostic and research tools for confirmation of late-presenting or asymptomatic MPXV cases, contact tracing, epidemiological studies, seroepidemiological surveys, and better understanding of the role of IgG and neutralizing antibodies in the immune response to infection and vaccination. A multidisciplinary approach combining advanced molecular techniques with traditional virological methods is important for rapid and reliable diagnosis, surveillance, and control of the outbreak.