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Introduction: Sclerosing pneumocytoma (SP) is a rare benign tumor and a potential diagnostic pitfall. Our aim was to review the cytologic features of our surgically diagnosed SP cases including the clinical, immunohistochemical and available molecular findings., Materials and Methods: A computerized search from 2013 to 2020 for surgical cases with corresponding cytology specimens diagnosed as SP was performed. The clinical data, cytology, and surgical specimens were collated for analysis., Results: Six cytology specimens were collected. All were female (mean age = 35). Three have incidental lung nodules and three with cough. Cytologic findings showed variable architectural pattern (papillary, solid, singly scattered, acinar/rosette-like) and cellular heterogeneity (surface, stromal, epithelioid, plasmacytoid cells). Atypia was identified in 4/6 cases. The original cytology diagnoses were negative = 1, SP = 2 and adenocarcinoma = 3. The latter diagnoses were amended to SP after review of the surgical specimens. The three false positive cases on review have cytologic features mimicking adenocarcinoma. Immunohistochemical stains showed tumor cells (surface and stromal) were positive for TTF-1, and EMA with only the surface cells positive for pancytokeratin and Napsin A. Though two cases sent for molecular testing were negative for AKT1 or CTNNB1 exon 3 mutation, our panel did not evaluate AKT1 exon 4., Conclusions: SP is a diagnostic pitfall with 50% initially misdiagnosed as adenocarcinoma. Integrating the clinical/radiologic findings, cytologic features, and performance of immunohistochemistry on cell block are helpful in avoiding misdiagnosis. Molecular testing for recurrent mutations, if present, could be helpful for diagnosis and possible therapy options. However, routinely used molecular testing may not always capture relevant molecular markers for SP., (© 2024 The Author(s). Diagnostic Cytopathology published by Wiley Periodicals LLC.)

Objectives: The rising rates of Helicobacter pylori antimicrobial resistance highlight the importance of better understanding its epidemiology in the pediatric population. We aim to study the epidemiological factors associated with antimicrobial resistance in a cohort of US children with H. pylori infection., Methods: A retrospective cohort study of patients in the New England region of the United States between January 15, 2015 and October 15, 2022, with a first-time diagnosis of H. pylori on gastric biopsy and who had antimicrobial resistance data available. Descriptive statistics and logistic regression models were used to determine associations between the patient's demographics, clinical factors, endoscopic findings, and antimicrobial resistance., Results: Out of 273 patients (46% male, median 12.8 years), 118 (43.2%) were resistant to at least one antimicrobial. Clarithromycin resistance (24.5%) was the highest, followed by metronidazole (21.6%), fluoroquinolones (9.9%), rifabutin (3.3%), and amoxicillin (2.6%). Clarithromycin resistance was higher in Asians compared to White individuals (odds ratio [OR]: 4.68, 95% confidence interval [CI]: [2.01-10.89], p < 0.001), and lower in Black compared to White individuals (OR: 0.23, 95% CI: [0.08-0.69], p = 0.01). Antimicrobial resistance to clarithromycin and metronidazole were similar among Hispanics. Asian individuals were more likely to demonstrate one or more antimicrobial resistances compared to White (OR: 3.66, 95% CI: [1.50-8.89], p < 0.001). Compared to individuals from higher household incomes (>$100,000), those from less than $30,000 household incomes had a higher risk of metronidazole resistance (OR: 7.94, 95% CI: [1.83-34.36], p = 0.01)., Conclusion: Our study provides novel information concerning the association of H. pylori antimicrobial resistance to race, socioeconomic status, and age in the pediatric population., (© 2024 European Society for Pediatric Gastroenterology, Hepatology, and Nutrition and North American Society for Pediatric Gastroenterology, Hepatology, and Nutrition.)

Background: Aetiological diagnosis and targeted antibiotic therapy are essential to improve the prognosis of patients with infective endocarditis. Molecular tests on blood have been reported to be effective in identifying the causative organism and are recommended when blood cultures are negative. The role of molecular tests on the surgically excised valve is still unclear and needs further investigation., Materials and Methods: In this prospective, observational, single center study, we enrolled 100 consecutive patients with native or prosthetic valve endocarditis who underwent cardiac surgery between April 2020 and June 2023. Results of preoperative blood cultures, valve culture, 16s ribosomal RNA and histopathologic analysis of surgical samples were collected in a dedicated database., Results: The mean age of the study population was 60 ± 12.5 years, with a majority of men (73%). Previous cardiac surgery was reported in 31% of patients. Blood culture, valve culture, and 16srRNA were positive in 83%, 47%, and 76% of cases, respectively. The sensitivity of both valve culture and 16srRNA decreased significantly with prolonged preoperative antibiotic therapy. Of note, 16srRNA was the only positive result in 7% of cases, allowing aetiological diagnosis. In 33% of patients, the valve culture test was negative while the molecular test was positive. In these cases, histopathological analysis showed acute inflammation in most cases. In 10%, the molecular test helped in resolving discrepancies between the results of blood and valve cultures., Conclusions: The molecular test showed significantly higher diagnostic sensitivity than valve culture and maintained this efficacy even after 28 days of preoperative antibiotic therapy. In addition to identifying the pathogen in 7% of cases with negative culture results, the molecular test demonstrated utility in other crucial situations. When valve cultures were negative, combining molecular testing and histopathological analysis they allowed the identification of patients who could benefit from prolonged antibiotic therapy. In addition, molecular testing guided the choice of antibiotic treatment when there was a discrepancy between blood culture and valve culture results. Based on these findings, molecular testing should be considered in all patients with infective endocarditis undergoing cardiac surgery. Clinical Trial Registration : ClinicalTrials.gov, identifier (NCT05791357)., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2024 Pavone, Cammertoni, Calabrese, Bruno, Scoppettuolo, Lombardo, Giovannenze, Taddei, Fiori, D'Inzeo, Cutrone, Iannaccone, Del Zanna and Massetti.)

Context: Molecular testing can refine the risk of malignancy in thyroid nodules with indeterminate cytology to decrease unnecessary diagnostic surgery., Objective: This study was performed to evaluate the outcomes of cytologically indeterminate thyroid nodules managed with Afirma genomic sequencing classifier (GSC) testing., Methods: Adult patients who underwent a biopsy at 3 major academic centers between July 2017 and June 2021 with Bethesda III or IV cytology were included. All patients had surgery or minimum follow-up of 1 year ultrasound surveillance. The primary outcomes were the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of GSC in Bethesda III and IV nodules., Results: The median nodule size of the 834 indeterminate nodules was 2.1 cm and the median follow-up was 23 months. GSC sensitivity, specificity, PPV, and NPV across all institutions were 95%, 81%, 50%, and 99% for Bethesda III nodules and 94%, 82%, 65%, and 98% for Bethesda IV nodules, respectively. The overall false-negative rate was 2%. The NPV of GSC in thyroid nodules with oncocytic predominance was 100% in Bethesda III nodules and 98% in Bethesda IV nodules. However, the PPV of oncocytic nodules was low (17% in Bethesda III nodules and 45% in Bethesda IV nodules). Only 22% of thyroid nodules with benign GSC results grew during surveillance., Conclusion: GSC is a key tool for managing patients with indeterminate cytology, including the higher-risk Bethesda IV category. GSC-benign thyroid nodules can be observed similarly to thyroid nodules with benign cytology., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Context: BRAF V600E mutation is the most common genetic driver of papillary thyroid cancer (PTC), where it is found with various allele frequency (AF), reflecting the proportion of cells carrying the mutant and wild-type gene alleles., Objective: To determine whether BRAF V600E AF can improve prognostication and inform initial surgical management of PTC., Design: Retrospective cohort study (2016-2019)., Setting: UCLA health., Patients: Consecutive patients with Bethesda V/VI nodules and isolated BRAF V600E mutation who underwent surgery with histopathology showing PTC., Interventions: Blinded ThyroSeq v3 molecular analysis after completion of initial management and follow-up., Main Outcomes Measures: Aggressive histopathology and cancer persistence/recurrence., Results: Of 73 patients, the median BRAF V600E AF was 25.5% (IQR, 16.7-34.3%). Higher median AF was seen in patients classified as American Thyroid Association (ATA) high-risk (37%) vs. intermediate-risk (25.3%, p<0.01) and low-risk (24.7%, p<0.01), largely attributed to higher AF in patients with gross extrathyroidal extension (ETE) (40.1% vs. 25.2% without gross ETE, p=0.02). No differences in AF were observed on the basis of lymph node positivity or presence of aggressive variants of PTC. A higher BRAF V600E AF was also found in patients with tumors ≥2cm vs. <2cm (median 32.0% vs. 24.4%, p<0.01). Over 4.1 years of follow-up, disease persistence/recurrence was found in 7 patients (9.4%) and was associated with higher median AF than those without recurrence (35.3% vs. 25.2%, p=0.02). Higher AF was associated with poorer recurrence-free survival (AF≥35%, HR 7.40, CI 1.4-38.1)., Conclusions: Higher AF was associated with gross ETE and increased recurrence risk. This may inform initial management in patients with PTC harboring an isolated BRAF V600E mutation., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com. See the journal About page for additional terms.)

Introduction: There is a lack of documentation regarding cytopathology of renal neuroendocrine neoplasms (NENs) due to their rarity., Materials and Methods: Five cytology cases were gathered from 3 institutes., Results: Cohort consisted of 4 females and 1 male. Fine needle aspiration biopsy and touch preparation slides of core needle biopsy revealed cellular samples, composed of round, plasmacytoid, or columnar cells. Tumor cells were present in nested, acinar, 3D cluster, and individual cell patterns. Tumor cells in 3 cases exhibited uniformly round to oval small nuclei with inconspicuous nucleoli, finely granular chromatin, and smooth nuclear membranes, whereas 2 other cases showed pleomorphic nuclei with conspicuous nucleoli, nuclear molding, and irregular nuclear membranes. Tumor cells displayed pale or granular cytoplasm, with 1 case showing small vacuoles. Examination of cores and cell blocks demonstrated tumor cells in sheets, nests, or acini. All tumor cells were positive for neuroendocrine immunomarkers. Based on mitotic count, Ki-67 index and morphology, 3 tumors were graded as well-differentiated neuroendocrine tumor (WDNET) (1 grade [G] 3, 1 G2, 1 G1) and 2 as large cell neuroendocrine carcinoma. Deletion of 7q, 10q, and 19q was detected in WDNETs. Two patients with large cell neuroendocrine carcinoma and 1 with WDNET G3 underwent chemotherapy due to aggressiveness, whereas nephrectomy was performed for patients with WDNET G1 and 2 without metastasis., Conclusions: Cytopathological characteristics of renal NENs closely resemble those affecting other organs. Despite its rarity, renal NENs should be kept in mind when confronted with morphological resemblances to NENs, to prevent misdiagnosis and inappropriate therapeutic interventions., (Copyright © 2024 American Society of Cytopathology. Published by Elsevier Inc. All rights reserved.)

Five cases of FISH verified BCOR rearranged high-grade endometrial stromal sarcoma were retrospectively analyzed. The patient age ranged from 33 to 65 years (median, 48.4 years). Most patients presented with irregular vaginal bleeding (3/5) and uterus mass (2/5). Only one patient developed an abdominal wall metastasis and other patients remained in good condition during the follow-up. Pathological findings revealed that the tumors exhibited morphological diversity in terms of cell shape, arrangement pattern and tumor stroma, compared to previous summarized histology of BCOR rearranged high-grade endometrial stromal sarcoma. Detailed description of such morphology changes expanded our understanding of the histology of BCOR rearranged high-grade endometrial stromal sarcoma. Due to the non-specificity of morphology in such malignancies, molecular testing is needed for confirmation in all patients., Competing Interests: None., (IJCEP Copyright © 2024.)

Background: Serological screening tests play a crucial role to diagnose gambiense human African trypanosomiasis (gHAT). Presently, they preselect individuals for microscopic confirmation, but in future "screen and treat" strategies they will identify individuals for treatment. Variability in reported specificities, the development of new rapid diagnostic tests (RDT) and the hypothesis that malaria infection may decrease RDT specificity led us to evaluate the specificity of 5 gHAT screening tests., Methods: During active screening, venous blood samples from 1095 individuals from Côte d'Ivoire and Guinea were tested consecutively with commercial (CATT, HAT Sero-K-SeT, Abbott Bioline HAT 2.0) and prototype (DCN HAT RDT, HAT Sero-K-SeT 2.0) gHAT screening tests and with a malaria RDT. Individuals with ≥ 1 positive gHAT screening test underwent microscopy and further immunological (trypanolysis with T.b. gambiense LiTat 1.3, 1.5 and 1.6; indirect ELISA/T.b. gambiense; T.b. gambiense inhibition ELISA with T.b. gambiense LiTat 1.3 and 1.5 VSG) and molecular reference laboratory tests (PCR TBRN3, 18S and TgsGP; SHERLOCK 18S Tids, 7SL Zoon, and TgsGP; Trypanozoon S 2 -RT-qPCR 18S2, 177T, GPI-PLC and TgsGP in multiplex; RT-qPCR DT8, DT9 and TgsGP in multiplex). Microscopic trypanosome detection confirmed gHAT, while other individuals were considered gHAT free. Differences in fractions between groups were assessed by Chi square and differences in specificity between 2 tests on the same individuals by McNemar., Results: One gHAT case was diagnosed. Overall test specificities (n = 1094) were: CATT 98.9% (95% CI: 98.1-99.4%); HAT Sero-K-SeT 86.7% (95% CI: 84.5-88.5%); Bioline HAT 2.0 82.1% (95% CI: 79.7-84.2%); DCN HAT RDT 78.2% (95% CI: 75.7-80.6%); and HAT Sero-K-SeT 2.0 78.4% (95% CI: 75.9-80.8%). In malaria positives, gHAT screening tests appeared less specific, but the difference was significant only in Guinea for Abbott Bioline HAT 2.0 (P = 0.03) and HAT Sero-K-Set 2.0 (P = 0.0006). The specificities of immunological and molecular laboratory tests in gHAT seropositives were 98.7-100% (n = 399) and 93.0-100% (n = 302), respectively. Among 44 reference laboratory test positives, only the confirmed gHAT patient and one screening test seropositive combined immunological and molecular reference laboratory test positivity., Conclusions: Although a minor effect of malaria cannot be excluded, gHAT RDT specificities are far below the 95% minimal specificity stipulated by the WHO target product profile for a simple diagnostic tool to identify individuals eligible for treatment. Unless specificity is improved, an RDT-based "screen and treat" strategy would result in massive overtreatment. In view of their inconsistent results, additional comparative evaluations of the diagnostic performance of reference laboratory tests are indicated for better identifying, among screening test positives, those at increased suspicion for gHAT., Trial Registration: The trial was retrospectively registered under NCT05466630 in clinicaltrials.gov on July 15 2022., (© 2024. The Author(s).)

The emergence of next-generation sequencing technologies and computational advances have expanded our understanding of gene expression regulation (i.e., the transcriptome). This has also led to an increased interest in using transcriptomic biomarkers to improve disease diagnosis and stratification, to assess prognosis and predict the response to treatment. Significant progress in identifying transcriptomic signatures for various clinical needs has been made, with large discovery studies accounting for challenges such as patient variability, unwanted batch effects, and data complexities; however, obstacles related to the technical aspects of cross-platform implementation still hinder the successful integration of transcriptomic technologies into standard diagnostic workflows. In this article, we discuss the challenges associated with integrating transcriptomic signatures derived using high-throughput technologies (such as RNA-sequencing) into clinical diagnostic tools using nucleic acid amplification (NAA) techniques. The novelty of the proposed approach lies in our aim to embed constraints related to cross-platform implementation in the process of signature discovery. These constraints could include technical limitations of amplification platform and chemistry, the maximal number of targets imposed by the chosen multiplexing strategy, and the genomic context of identified RNA biomarkers. Finally, we propose to build a computational framework that would integrate these constraints in combination with existing statistical and machine learning models used for signature identification. We envision that this could accelerate the integration of RNA signatures discovered by high-throughput technologies into NAA-based approaches suitable for clinical applications., Competing Interests: Declaration of interests This article is the subject of a patent application filed by Imperial College London, Patent Application Publication No. WO2024023491A1, “A method to optimise transcriptomic signatures”, where JRM, DHC, HRJ, LM and MK are named inventors. All other declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Viral respiratory infections represent a major threat to the population's health globally. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes COVID-19 disease and in some cases the symptoms can be confused with Influenza disease caused by the Influenza A viruses. A simple, fast, and selective assay capable of identifying the etiological agent and differentiating the diseases is essential to provide the correct clinical management to the patient. Herein, we described the development of a genomagnetic assay for the selective capture of viral RNA from SARS-CoV-2 and Influenza A viruses in saliva samples and employing a simple disposable electrochemical device for gene detection and quantification. The proposed method showed excellent performance detecting RNA of SARS-CoV-2 and Influenza A viruses, with a limit of detection (LoD) and limit of quantification (LoQ) of 5.0 fmol L -1 and 8.6 fmol L -1 for SARS-CoV-2, and 1.0 fmol L -1 and 108.9 fmol L -1 for Influenza, respectively. The genomagnetic assay was employed to evaluate the presence of the viruses in 36 saliva samples and the results presented similar responses to those obtained by the real-time reverse transcription-polymerase chain reaction (RT-PCR), demonstrating the reliability and capability of a method as an alternative for the diagnosis of COVID-19 and Influenza with point-of-care capabilities., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Precise classification of sarcomas is crucial to optimal clinical management. In this prospective, multicenter, observational study within the Hellenic Group of Sarcoma and Rare Cancers (HGSRC), we assessed the effect of expert pathology review, coupled with the application of molecular diagnostics, on the diagnosis and management of sarcoma patients. Newly diagnosed sarcoma patients were addressed by their physicians to one of the two sarcoma pathologists of HGSRC for histopathological diagnostic assessment. RNA next-generation sequencing was performed on all samples using a platform targeting 86 sarcoma gene fusions. Additional molecular methods were performed in the opinion of the expert pathologist. Therefore, the expert pathologist provided a final diagnosis based on the histopathological findings and, when necessary, molecular tests. In total, 128 specimens from 122 patients were assessed. Among the 119 cases in which there was a preliminary diagnosis by a non-sarcoma pathologist, there were 37 modifications in diagnosis (31.1%) by the sarcoma pathologist, resulting in 17 (14.2%) modifications in management. Among the 110 cases in which molecular tests were performed, there were 29 modifications in diagnosis (26.4%) through the genomic results, resulting in 12 (10.9%) modifications in management. Our study confirms that expert pathology review is of utmost importance for optimal sarcoma diagnosis and management and should be assisted by molecular methods in selected cases.

In ICU settings, screening patients upon admission for potential multiresistant bacteria (BMR) carriers is crucial. Traditionally, clinical decisions relied on delayed culture results, but a rapid PCR molecular test called RealCycler-Rezero-U/G (Progenie-molecular©), emerged as an alternative. This study aimed to validate its effectiveness in detecting gram-negative BMR in rectal swabs at ICU admission. Over 24 months, an observational study was conducted on 1,234 admitted patients, with 217 meeting isolation criteria and undergoing both PCR and culture tests. Results showed a 17.5 % positive rate for screening. The PCR test exhibited impressive accuracy at 98.6 % and a strong negative predictive value of 99.4 %. The area under the ROC curve (AUC) was 0.98, indicating high reliability. Notably, PCR results were available 44.5 h earlier than culture. In conclusion, PCR-based molecular testing for gram-negative BMR offers excellent diagnostic performance and a valuable negative predictive value, making it a suitable screening tool for ICU admissions., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

With the continuous spread of new SARS-CoV-2 variants of concern (VOCs), the monitoring of diagnostic test performances is mandatory. We evaluated the changes in antigen diagnostic tests' (ADTs) accuracy along the Delta to Omicron VOCs transition, exploring the N protein mutations possibly affecting ADT sensitivity and assessing the best sampling site for the diagnosis of Omicron infections. In total, 5175 subjects were enrolled from 1 October 2021 to 15 July 2022. The inclusion criteria were SARS-CoV-2 ADT combined with a same-day RT-PCR swab test. For the sampling site analysis, 61 patients were prospectively recruited during the Omicron period for nasal and oral swab analyses by RT-PCR. Next-Generation Sequencing data were obtained to evaluate the different sublineages. Using RT-PCR as a reference, 387 subjects resulted in becoming infected and the overall sensitivity of the ADT decreased from 63% in the Delta period to 33% in the Omicron period. This decrease was highly statistically significant ( p < 0.001), and no decrease in viral load was detected at the RNA level. The nasal site presented a significantly higher viral load than the oral site during the Omicron wave. The reduced detection rate of Omicron infections by ADT should be considered in the global testing strategy to preserve accurate diagnoses across the changing SARS-CoV-2 variants.

Background: The current preoperative malignancy risk evaluation for thyroid nodules involves stepwise diagnostic modalities including ultrasonography, thyroid function serology and fine-needle aspiration (FNA) cytopathology, respectively. We aimed to substantiate the stepwise contributions of each diagnostic step and additionally investigate the diagnostic significance of quantitative chromogenic imprinted gene in-situ hybridization (QCIGISH)-an adjunctive molecular test based on epigenetic imprinting alterations., Methods: A total of 114 cytopathologically-diagnosed and histopathologically-confirmed thyroid nodules with complete ultrasonographic and serological examination records were evaluated using QCIGISH in the study. Logistic regression models for thyroid malignancy prediction were developed with the stepwise addition of each diagnostic modality and the contribution of each step evaluated in terms of discrimination performance and goodness-of-fit., Results: From the baseline model using ultrasonography [area under the receiver operating characteristics curve (AUROC): 0.79; 95% confidence interval (CI): 0.71-0.86], significant improvements in thyroid malignancy discrimination were observed with the stepwise addition of thyroid function serology (AUROC: 0.82; 95% CI: 0.74-0.90; P=0.23) and FNA cytopathology (AUROC: 0.88; 95% CI: 0.81-0.94; P=0.02), respectively. The inclusion of QCIGISH as an adjunctive molecular test further advanced the preceding model's diagnostic performance (AUROC: 0.95; 95% CI: 0.91-1.00, P=0.007)., Conclusions: Our study demonstrated the significant stepwise diagnostic contributions of standard clinical assessments in the malignancy risk stratification of thyroid nodules. However, the addition of molecular imprinting detection further enabled a more accurate and definitive preoperative evaluation especially for morphologically indeterminate thyroid nodules and cases with potentially discordant results among standard modalities.

Inappropriate ordering practices, either under or over ordering of diagnostic tests, are recognized problems with possible negative downstream consequences. As the menu of clinical tests, especially molecular tests grows, it is becoming increasingly important to provide guidance to providers on the appropriate utilization. Diagnostic stewardship programs have been established at many institutions to help direct the appropriate utilization of laboratory testing to ultimately guide patient management and treatment decisions. Many molecular tests have now received Clinical Laboratory Improvement Amendments (CLIA)-waived status for use in a point-of-care (POC) setting; however, parallel diagnostic stewardship programs have not been established to help guide providers on how best to use these tests. In this article, we will discuss the available molecular POC tests and opportunities and challenges for establishing diagnostic stewardship programs for molecular testing performed in the POC setting., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Brucellosis is a significant infection that causes abortion, decreased milk production, and sterility in livestock, which greatly affects the industry. This study aimed to determine the prevalence of Brucella in buffalo milk samples across various regions of Iran, utilizing serological, molecular, and cultural analyses. A total of 1860 buffalo milk samples were collected from industrial, semi-industrial, and traditional buffalo farms in four major buffalo breeding provinces. The milk ring test agglutination test (MRT) was initially conducted on all milk samples, followed by culture and molecular testing for positive and negative samples in MRT. The study revealed positive results for the presence of Brucella DNA in various provinces of Iran. The MRT had a relatively low sensitivity, with results ranging from 0 to 0.7% in different provinces. However, the AMOS PCR method showed a significantly higher presence of Brucella DNA, ranging from 13 to 46% in these provinces. The highest abundance of Brucella bacterial DNA was found in Ardabil province, while the lowest was in West Azerbaijan province. Brucella abortus was the most commonly detected bacteria, followed by Brucella melitensis. Interestingly, the B. abortus vaccine strain RB51 was detected in 26.3% of positive samples of B. abortus. The culture assay of milk samples further confirmed the presence of B. melitensis biovar 1 in one sample from Khuzestan province. Overall, the study emphasizes that the AMOS PCR method is the most sensitive in detecting Brucella-exposed milk, while the sensitivity of milk sample culture and MRT is relatively lower., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

Background: We assessed the laboratory diagnosis and treatment of invasive fungal disease (IFD) in Italy to detect limitations and potential for improvement., Methods: The survey was available online at www.clinicalsurveys.net/uc/IFI management capacity/, and collected variables such as (a) institution profile, (b) perceptions of IFD in the respective institution, (c) microscopy, (d) culture and fungal identification, (e) serology, (f) antigen detection, (g) molecular tests, (h) susceptibility testing and (i) therapeutic drug monitoring (TDM)., Results: The laboratory capacity study received responses from 49 Italian centres, with an equitable geographical distribution of locations. The majority of respondents (n = 36, 73%) assessed the occurrence of IFD as moderate-high, with Aspergillus spp. being the pathogen of highest concern, followed by Candida spp. and Mucorales. Although 46 (94%) of the institutions had access to microscopy, less than half of them performed direct microscopy on clinical specimens always when IFD was suspected. Cultures were available in all assessed laboratories, while molecular testing and serology were available in 41 (83%), each. Antigen detection tests and antifungal drugs were also generally accessible (> 90%) among the participating institutions. Nevertheless, access to TDM was limited (n = 31, 63%), with a significant association established between therapeutic drug monitoring availability and higher gross domestic product per capita., Conclusions: Apart from TDM, Italy is adequately prepared for the diagnosis and treatment of IFD, with no significant disparities depending on gross domestic product. Future efforts may need to focus on enhancing the availability and application of direct microscopic methods, as well as TDM, to promote optimal treatment and better patient outcomes., (© 2023. The Author(s).)

Background: Cytologically indeterminate thyroid nodules (ITN) pose a management challenge. Here we analyze if adding ultrasound characteristics to Afirma Genome Sequence Classifier (GSC) results increases GSC diagnostic performance., Methods: We retrospectively analyzed 237 GSC-tested Bethesda III/IV ITNs between July 2017 and December 2019 and classified them by American Thyroid Association (ATA) and the Thyroid Imaging Reporting and Data System (TIRADS) of the American College of Radiology., Results: The benign call rate was higher in Bethesda III ITNs with TIRADS <5 vs TIRADS 5 (89% vs 68%. P = .015). The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of GSC in ATA high-risk Bethesda III ITNs vs lower were 100% vs 80% ( P = 1), 89.5% vs 91.5% ( P = .67), 66.7% vs 25% ( P = .13), and 100% vs 99.2% ( P = 1), respectively, and for TIRADS 5 vs <5 were 100% vs 80% ( P = 1), 88.2% vs 91.4% ( P = .65), 71.4% vs 23.5% ( P = .06), and 100% vs 99.3% ( P = 1). The sensitivity, specificity, PPV, and NPV of GSC in high-risk ATA Bethesda IV ITNs vs lower were 66.7% vs 100% ( P = .42), 83.3% vs 85.7% ( P = 1), 66.7% vs 64.3% ( P = 1), and 83.3% vs 100% ( P = .3), respectively, and for TIRADS 5 vs <5 were 66.7% vs 90% ( P = .42), 88.9% vs 83.8% ( P = 1), 66.7% vs 60% ( P = 1), and 88.9% vs 96.9% ( P = .39)., Conclusion: Sensitivity, specificity, NPV, and PPV of GSC were not significantly different in ATA high-risk and TIRADS 5 ITNs compared to ATA < high-risk and TIRADS 1-4 ITNs., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society.)

Background: Antifungal diagnostic capacity has been documented in various countries, there is a lack of comprehensive research on clinical mycology diagnostics and treatment in Hungary., Methods: We conducted an online survey encompassing questions that explored various aspects of the mycology diagnostic and antifungal therapy-related information. The survey aimed to gather details about institutional profiles, perceptions of invasive fungal infections (IFIs), and access to microscopy, culture, serology, antigen detection, molecular testing, and therapeutic drug monitoring., Results: As of May 2023, a total of 17 institutions responded to the questionnaire. Seven participants categorized the institutional incidence of IFI as 'very low', four as 'low', and six as 'mild'. The majority of centers identified Candida spp. (94%) and Aspergillus spp. (82%) as the most prevalent fungal pathogens. Nearly half of the laboratories (47%) reported using matrix-assisted laser desorption/ionization-time of flight mass spectrometry for identification. All institutions had access to microscopy and culture-based diagnostic approaches. A significant number of centers had access to antigen detection (71%) and various molecular assays (59%). Regarding antifungal agents, all reporting sites used at least one triazole, with voriconazole (77%) being the most common mold-active azole. Furthermore, 71% of the centers applied at least one formulation of amphotericin B, and 65% to one echinocandin. However, only 18% of the centers used 5-flucytosine., Conclusion: Resource availability for diagnosing and treating IFI in Hungary varies across hospitals based on location. Surveys help identify gaps and limitations in this area. To address these challenges, interregional cooperation within Hungary could be a facilitating strategy., Competing Interests: RK has no conflict of interest. LM has received conference travel grants from MSD, Astellas, and Pfizer. JS has received research support from the Ministry of Education and Research (BMBF) and Basilea Pharmaceuticals Inc., outside the submitted work; has received speaker honoraria from Pfizer Inc., Gilead, and AbbVie, outside the submitted work; has been a consultant to Gilead, Produkt&Markt GmbH, Alvea Vax. and Micron Research, outside the submitted work; and has received travel grants by German Society for Infectious Diseases (DGI e.V.) and Meta-Alexander Foundation, outside the submitted work. OAC reports grants or contracts from Amplyx, Basilea, BMBF, Cidara, DZIF, EU-DG RTD (101037867), F2G, Gilead, Matinas, MedPace, MSD, Mundipharma, Octapharma, Pfizer, Scynexis; Consulting fees from Amplyx, Biocon, Biosys, Cidara, Da Volterra, Gilead, Matinas, MedPace, Menarini, Molecular Partners, MSG-ERC, Noxxon, Octapharma, PSI, Scynexis, Seres; Honoraria for lectures from Abbott, Al-Jazeera Pharmaceuticals, Astellas, Grupo Biotoscana/United Medical/Knight, Hikma, MedScape, MedUpdate, Merck/MSD, Mylan, Pfizer; Payment for expert testimony from Cidara; Participation on a Data Safety Monitoring Board or Advisory Board from Actelion, Allecra, Cidara, Entasis, IQVIA, Janssen, MedPace, Paratek, PSI, Shionogi; a patent at the German Patent and Trade Mark Office (DE 10 2021 113 007.7), outside of the submitted work. JSG reports speaker honoraria from Gilead and Pfizer, outside of the submitted work., (© The Author(s), 2023.)

Background: The diagnosis of extrapulmonary tuberculosis (EPTB) shows numerous difficulties because of non-specific symptomatology and low sensitivity of conventional methods. Loop-mediated isothermal amplification (LAMP) is a fast and low-cost technique, which can amplify under isothermal conditions an amount of target DNA copies into approximately a billion copies., Objective: The present study aimed to evaluate a IS6110-LAMP system for Mycobacterium tuberculosis detection in blood and urine samples from patients with EPTB., Methods: The collected samples (n = 122) were stratified in two groups: Group EPTB - patient samples with confirmed EPTB (n = 61); Group non-TB - patient samples without TB (n = 61). The urine samples underwent decontamination, and the components of blood samples were separated (plasma and PBMC). DNA extractions were performed in all biological samples followed by IS6110-LAMP assay technique. The detection limit was evaluated through dilution curves (1:10) using Mtb reference strain (H37Rv) genomic DNA., Findings: The detection limit of IS6110-LAMP was 10 fg/μL (∼10-20 bacilli/μL). The IS6110-LAMP technique sensitivity and specificity were 95.65 % and 79.25 %, respectively, with a general kappa agreement index of 0.762., Main Conclusions: Based on these results, IS6110-LAMP test showed considerable diagnostic parameters, being able to aid in the speed and accuracy of the final EPTB diagnosis., Competing Interests: Conflict of interest The authors declared no conflicts of interest., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Background: Staphylococcus aureus is a typical pathogenic agent that causes several morbidities and mortalities which variable largely following the severity of bacteria and activity of host immunity., Aim: Isolation of S. aureus from different cattle infections, molecular detection of nuc gene in positive S. aureus isolates, and identification of the effectiveness of the phagocytic activity., Methods: Totally, 100 cattle with various infections (25 wounds, 25 abscesses, 25 nasal discharges, and 25 ear swaps) were selected and subjected to collection of swabs under controlled conditions. All collected samples were cultured on mannitol salt agar (MSA) and assessed by biochemical tests. Targeting the nuc gene, all study MSA positive isolates were examined molecularly by conventional polymerase chain reaction (PCR), and then subjected to antibiotic susceptibility test. Jugular venous blood was collected from all infected animals in addition to 20 healthy cattle that were selected as a control group to estimate the phagocytic activity of S. aureus isolates., Results: The findings of MSA culture revealed a total of 80 positive samples of S. aureus as 23, 21, 20, and 16 positive isolates for nasal discharge, abscess, wound, and ear swab, respectively; based on its morphology, cultural trait, and biochemical test. Subsequently, PCR assaying of MSA-positive isolates demonstrated an overall 59 positive samples as 14, 16, 12, and 17 positive isolates for nasal discharge, abscess, wound, and ear swabs, respectively. Antibiotic susceptibility testing of S. aureus -positive PCR isolates reported a significantly high sensitivity to chloramphenicol and vancomycin, and a high resistance to penicillin. Finally, there was a considerable decline in phagocytic activity in particular 2 weeks post-infection as a result of bacterial invasion., Conclusion: This study shows a high prevalence of S. aureus in cattle infections, and the protocol includes a regular screening of cattle infection and suitable therapy based on antibacterial susceptibility test is of great importance in the long-term control of the pathogen. However, additional molecular studies targeting other genes of S. aureus and the role of immune markers in different infections should be aimed., Competing Interests: The authors declare that there is no conflict of interest

Purpose: To evaluate the diagnostic efficacy of CapitalBio Mycobacterium real-time polymerase chain reaction assay (CapitalBio test) in spinal tuberculosis (STB). The value of histopathology combined with the CapitalBio test in diagnosing STB was also assessed., Methods: We retrospectively analyzed the medical information of suspected STB. The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and area under the curve (AUC) of histopathology, CapitalBio test, and histopathology combined with CapitalBio test were calculated to evaluate their diagnostic efficacy compared with a composite reference standard., Results: A total of 222 suspected STB patients were included in the study. The sensitivity, specificity, PPV, NPV, and AUC of histopathology for STB were recorded to be 62.0, 98.0, 97.4, 68.3%, and 0.80, respectively. The sensitivity, specificity, PPV, NPV, and AUC of the CapitalBio test were 75.2, 98.0, 97.9, 76.7%, and 0.87, respectively, while that of histopathology combined with the CapitalBio test was 81.0, 96.0, 96.1, 80.8%, and 0.89, respectively., Conclusion: Histopathology and CapitalBio test exhibited high accuracy and are recommended in diagnosing STB. Histopathology combined with the CapitalBio test might give the best efficacy in STB diagnosis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Yao, Bu, Zhang and Zhang.)

Background: Frequent serial monitoring of plasma cytomegalovirus (CMV) viral load caused unnecessary budgets for laboratory testing without changes in treatment. We aimed to implement diagnostic stewardship to limit CMV viral load testing at appropriate intervals., Methods: A quasi-experimental study was performed. To avoid unnecessary plasma CMV viral load testing, the inpatient electronic pop-up reminder was launched in 2021. In cases with plasma CMV viral load testing was ordered in intervals of less than five days, telephone interview and feedback were performed. Pre-post intervention data was compared in terms of clinical and monetary outcomes. The rate of plasma CMV viral load testing performed in intervals of less than five days was compared between 2021 and 2019 using the Poisson regression model., Results: After the protocol implementation, there was a significant decrease in the rate of plasma CMV viral load test orders in intervals of less than five days from 17.5% to 8.0% [incidence rate ratio 0.40, p < 0.001]. There was no statistically significant difference in the incidence of CMV DNAemia and CMV disease (p = 0.407 and 0.602, respectively). As a result, the hospital could save the costs of plasma CMV viral load testing per 1,000 patients performed with intervals of less than five days from 2,646,048.11 to 1,360,062.89 Thai Baht., Conclusions: The diagnostic stewardship program is safe and helpful in reducing unnecessary plasma CMV viral load testing and costs., (© 2023. The Author(s).)

Immunosuppressed patients, transplant recipients, and those with acute or chronic respiratory disease are at increased risk for invasive fungal infections in Argentina. Although the national public system guarantees universal access to health care for all citizens, little is known about the quality of available diagnostic and treatment armamentaria for invasive fungal infections in the country. Between June and August 2022, infectious disease clinicians from each of the 23 provinces and the Autonomous City of Buenos Aires were contacted to describe local access to fungal diagnostic tools and antifungal agents. The information collected included different aspects such as hospital characteristics, patients admitted and wards, access to diagnostic tools, estimated infection incidence, and treatment capacity. Thirty responses were collected from facilities throughout Argentina. Most institutions were governmental (77%). A mycology department was available in 83% of them. Histopathology was available in almost 93% of the sites, while automated methods and galactomannan tests were available in 57%, each; 53% of the sites had access to MALDI-TOF-MS through regional reference laboratories, and PCR was present in 20% of the sites. Susceptibility testing was available in 63% of the laboratories. Candida spp. (24%), Cryptococcus spp. (20%), Aspergillus spp. (18%), and Histoplasma spp. (16%) were described as the main pathogens. Fluconazole was the only antifungal agent available in all institutions. This was followed by amphotericin B deoxycholate (83%) and itraconazole (80%). If an antifungal agent was not available onsite, then 60% of the patients could receive adequate antifungal treatment within the first 48 h upon request. Although there are no significant differences in access to diagnostic and clinical management of invasive fungal infections among the Argentinean centres studied, national awareness-raising initiatives led by policymakers could help to improve their general availability., (© The Author(s) 2023. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology.)

Purpose: Fine-needle aspiration (FNA) biopsy is an effective method to discriminate malignant thyroid nodules but reaches indeterminate results in approximately 30% of cases. Molecular testing can improve the diagnostic accuracy of FNA. This study aimed to investigate the real-life utility of the five-gene panel testing in thyroid FNAs., Methods: 759 thyroid nodules from 740 patients under FNAs were retrospectively enrolled. Gene mutation information and clinical parameters, including age, gender, tumor size, and lymph node metastasis, were respectively recorded. Cytological results were classified based on The Bethesda System for Reporting Thyroid Cytopathology (TBSRTC). We analyzed mutational hotspots in BRAF, KRAS, NRAS, HRAS, and TERT genes from FNA specimens. The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and accuracy were calculated to assess diagnostic performance., Results: We identified 549 five-gene mutations in 759 nodules (72.3%), and the mutation frequency increased from the lower TBSRTC category to the upper category. BRAF.p.V600E showed the highest mutation incidence (71.3%) in the five-gene panel, correlated with the small to medium diameter (p = 0.008, p = 0.012) and high cytological categories (p < 0.001). The sensitivity, specificity, PPV, NPV, and accuracy of the combination of FNA cytology and five-gene detection were 96.83%, 100%, 100%, 42.86%, and 96.90%, respectively., Conclusions: The mutation frequency of the five-gene panel is 72.3% in thyroid FNAs. BRAF.p.V600E has the highest alteration rate, which is closely associated with tumor size and cytological results. The five-gene panel can improve the sensitivity and accuracy of FNA cytology, which may represent a valid adjunct technique in distinguishing thyroid nodules., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Purpose: To evaluate the accuracy of metagenomic next-generation sequencing (mNGS) for rapid diagnosis of spinal tuberculosis using spinal tissue specimens., Methods: Medical data regarding suspected spinal tuberculosis were retrospectively analyzed. The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and area under the curve (AUC) of mNGS were calculated to determine its diagnostic efficacy, and these values were compared with culture and composite reference standard (CRS)., Results: Two hundred and three cases of spinal tuberculosis were included for analysis. The sensitivity, specificity, PPV, NPV, and AUC of mNGS compared with culture were all very good. When CRS was used for the comparison, the sensitivity of mNGS and culture were 71.2% and 73.0%; the specificity and PPV were 100% in all cases; the NPV were 74.2% and 75.4%; the AUCs were all 0.86. The sensitivity and NPV of culture were slightly higher than that of mNGS; however, the diagnostic efficacy of mNGS and culture was consistent (P > 0.05)., Conclusion: Spinal tissue specimens for mNGS testing had very good accuracy for diagnosing spinal tuberculosis., Competing Interests: The authors declare that there is no conflict of interest., (© 2023 Jin et al.)

Background and Aim: Emergence of drug resistance, especially to second-line drugs, hampers tuberculosis elimination efforts. The present study aimed to evaluate MTBDRplus and MTBDRsl assays for detecting first-line and second-line drug resistance, respectively, in gastrointestinal tuberculosis (GITB)., Methods: Thirty ileocecal biopsy specimens, processed in the Department of Microbiology between 2012 and 2022, that showed growth of Mycobacterium tuberculosis on culture were included in the study. DNA, extracted from culture, was subjected to MTBDRplus and MTBDRsl (Hain Lifescience GmbH, Nehren, Germany), following manufacturer's instructions. Their performance was compared against phenotypic drug susceptibility testing (pDST) and gene sequencing., Results: Out of the 30 specimens, 4 (13.33%) were mono-isoniazid resistant, 4 (13.33%) were multidrug resistant (MDR), 2 (6.67%) were pre-extensively drug resistant (pre-XDR), and 2 (6.67%) were mono-fluoroquinolone resistant. The results were 100% concordant with pDST and gene sequencing., Conclusions: In the wake of growing drug resistance in all forms of extrapulmonary tuberculosis, including GITB, MTBDRplus and MTBDRsl are reliable tools for screening of resistance to both first-line and second-line drugs., (© 2023 Journal of Gastroenterology and Hepatology Foundation and John Wiley & Sons Australia, Ltd.)

Objective: Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant syndrome characterized by its clinical variability and complexity in diagnosis and treatment. We performed both clinical and molecular descriptions of four families with MEN1 in a follow-up at a tertiary center in Brasília., Methods: From a preliminary review of approximately 500 medical records of patients with pituitary neuroendocrine tumor (PitNET) from the database of the Neuroendocrinology Outpatient Clinic of the University Hospital of Brasília, a total of 135 patients met the criteria of at least two affected family members. From this cohort, we have identified 34 families: only four with a phenotype of MEN1 and the other 30 families with the phenotype of familial isolated pituitary adenoma (FIPA). Eleven patients with a clinical diagnosis of MEN1 from these four families were selected., Results: Variants in MEN1 gene were identified in all families. One individual from each family underwent genetic testing using targeted high-throughput sequencing (HTS). All patients had primary hyperparathyroidism (PHPT), and the second most common manifestation was PitNET. One individual had well-differentiated liposarcoma, which has been previously reported in a single case of MEN1. Three variants previously described in the database and a novel variant in exon 2 have been found., Conclusions: The study allowed the genotypic and phenotypic characterization of families with MEN1 in a follow-up at a tertiary center in Brasília., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Miranda, Valadares, Barra, Mesquita, de Santana, de Castro, Rita and Naves.)

Background: Molecular and antigen point-of-care tests (POCTs) have augmented our ability to rapidly identify and manage SARS-CoV-2 infection. However, their clinical performance varies among individual studies., Objectives: The evaluation of the performance of molecular and antigen-based POCTs in confirmed, suspected, or probable COVID-19 cases compared with that of laboratory-based RT-PCR in real-life settings., Data Sources: MEDLINE/PubMed, Scopus, Embase, Web of Science, Cochrane Library, Cochrane COVID-19 study register, and COVID-19 Living Evidence Database from the University of Bern., Study Eligibility Criteria: Peer-reviewed or preprint observational studies or randomized controlled trials that evaluated any type of commercially available antigen and/or molecular POCTs for SARS-CoV-2, including multiplex PCR panels, approved by the United States Food and Drug Administration, with Emergency Use Authorization, and/or marked with Conformitè Europëenne from European Commission/European Union., Participants: Close contacts and/or patients with symptomatic and/or asymptomatic confirmed, suspected, or probable COVID-19 infection of any age., Test/s: Molecular and/or antigen-based SARS-CoV-2 POCTs., Reference Standard: Laboratory-based SARS-CoV-2 RT-PCR., Assessment of Risk of Bias: Eligible studies were subjected to quality-control and risk-of-bias assessment using the Quality Assessment of Diagnostic Accuracy Studies 2 tool., Methods of Data Synthesis: Summary sensitivities and specificities with their 95% CIs were estimated using a bivariate model. Subgroup analysis was performed when at least three studies informed the outcome., Results: A total of 123 eligible publications (97 and 26 studies assessing antigen-based and molecular POCTs, respectively) were retrieved from 4674 initial records. The pooled sensitivity and specificity for 13 molecular-based POCTs were 92.8% (95% CI, 88.9-95.4%) and 97.6% (95% CI, 96.6-98.3%), respectively. The sensitivity of antigen-based POCTs pooled from 138 individual evaluations was considerably lower than that of molecular POCTs; the pooled sensitivity and specificity rates were 70.6% (95% CI, 67.2-73.8%) and 98.9% (95% CI, 98.5-99.2%), respectively., Discussion: Further studies are needed to evaluate the performance of molecular and antigen-based POCTs in underrepresented patient subgroups and different respiratory samples., (Copyright © 2022 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Ebola is a highly pathogenic virus, which in humans reaches a mortality rate above 50%. Due to a lack of laboratories in territories where Ebola viruses are endemic and the limited number of surveillance programmes, tests for the confirmation of suspected cases of Ebola are often performed in Reference Laboratories. While this provides guarantees regarding the accuracy of results, the shipment of samples to a centralized facility where the diagnostic test can be performed and the time required to achieve the results takes several days, which increases costs and entails delays in the isolation of positive subjects and therapeutic intervention with negative consequences both for patients and the community. Molecular tests have been the most frequently used tool in Ebola diagnosis in recent outbreaks. One of the most commonly used molecular tests is the Real-Star Altona, which targets a conserved area of the L gene. This assay showed different sensitivities depending on the Ebola virus: 471 copies/mL (EBOV) and 2871 copies/ml (SUDAN virus). The Cepheid system also showed good sensitivity (232 copies/mL). The LAMP platform is very promising because, being an isothermal reaction, it does not require high-precision instrumentation and can be considered a Point of Care (PoC) tool. Its analytical sensitivity is 1 copy/reaction. However, since data from real life studies are not yet available, it is premature to give any indications on its feasibility. Moreover, in November 2014, the WHO recommended the development of rapid diagnostic tests (RDT) according to ASSURED criteria. Several RDT assays have since been produced, most of which are rapid tests based on the search for antibody anti-Ebola viral proteins with immunochromatographic methods. Several viral antigens are used for this purpose: VP40, NP and GP. These assays show different sensitivities according to the protein used: VP40 57.4-93.1%, GP 53-88.9% and 85% for NP compared to reference molecular assays. From these results, it can be deduced that no RDT reaches the 99% sensitivity recommended by the WHO and therefore any RDT negative results in suspected cases should be confirmed with a molecular test., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Bettini, Lapa and Garbuglia.)

We performed a prospective study to determine if the pretest probability of a positive loop-mediated isothermal amplification test is greater when there are more signs and symptoms of GAS pharyngitis. Patients were enrolled if a clinician obtained a GAS RADT. The McIsaac score was calculated. The prevalence of positive LAMP and RADT results increased as the McIsaac score increased. The calculated sensitivity of LAMP was superior to RADT., (© The Author(s) 2022. Published by Oxford University Press on behalf of The Journal of the Pediatric Infectious Diseases Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Lung cancer remains the leading cause of cancer deaths worldwide. International societies have promoted the molecular analysis of MET proto-oncogene, receptor tyrosine kinase ( MET ) exon 14 skipping for the clinical stratification of non-small cell lung cancer (NSCLC) patients. Different technical approaches are available to detect MET exon 14 skipping in routine practice. Here, the technical performance and reproducibility of testing strategies for MET exon 14 skipping carried out in various centers were evaluated. In this retrospective study, each institution received a set ( n = 10) of a customized artificial formalin-fixed paraffin-embedded (FFPE) cell line (Custom MET ex14 skipping FFPE block) that harbored the MET exon 14 skipping mutation (Seracare Life Sciences, Milford, MA, USA), which was previously validated by the Predictive Molecular Pathology Laboratory at the University of Naples Federico II. Each participating institution managed the reference slides according to their internal routine workflow. MET exon 14 skipping was successfully detected by all participating institutions. Molecular analysis highlighted a median Cq cut off of 29.3 (ranging from 27.1 to 30.7) and 2514 (ranging from 160 to 7526) read counts for real-time polymerase chain reaction (RT-PCR) and NGS-based analyses, respectively. Artificial reference slides were a valid tool to harmonize technical workflows in the evaluation of MET exon 14 skipping molecular alterations in routine practice.

Introduction: DNA extracted from cytologic samples is occasionally used for various molecular tests. The aim of this study was to evaluate DNA extracted from differently prepared cytologic slides that can be used for PCR-based molecular tests., Methods: For each 23 cases of papillary thyroid carcinoma or colorectal adenocarcinoma tissues, six touch-imprinted cytological slides were prepared (group 1∼3), and remnant tissues were blocked for FFPE tissue (group 4). Cytologic slides were grouped by preparation methods: air-dried slides (group 1), fixed slides (group 2), and stained slides (group 3). Fixed slides were classified as 95% ethanol fixed (group 2A) and CytoRich Red Preservative solution fixed (group 2B). Stained slides were divided in 3 ways: Giemsa, Pap, and H&E stained (group 3A, 3B, and 3C, respectively). DNA extracted from each group was evaluated for concentration, 260/280 ratio, DNA Integrity Number (DIN) value, and mutation., Results: DNA concentration was highest in group 1 and lowest in group 2B. DIN value was highest in group 2A and lowest in group 2B. A mutation of BRAF or KRAS genes was detected in 18 FFPE tissue samples. Matched DNA extracts from groups 1, 2A, and 3 produced results consistent with FFPE tissue results, while mutation testing was successful for only four samples of DNA from group 2B., Conclusion: The mutation tests worked well for most samples except CytoRich Red Preservative-fixed slides. This study indicates that stained and unstained cytologic slides are a suitable source of PCR-based molecular tests as long as they are fixed in ethanol or stored for a short time in an air-dried condition., (© 2022 The Author(s). Published by S. Karger AG, Basel.)

The present study examined and reported Hepatocystis sp. infection in wild-caught vervet monkeys imported from Tanzania into the Razi vaccine and serum research institute (RVSRI). Polymerase chain reaction (PCR) targeting of 18S rRNA gene, followed by sequencing, Basic Local Alignment Search Tool (BLAST), and phylogenetic studies revealed that 82.8% of the imported monkeys were infected with Hepatocystis . Nevertheless, as illustrated by a routine parasitological examination of blood smears and histopathological examination of liver collected samples, the rates of Hepatocystis infection were obtained at 33.9% and 38.1%, respectively. Two isolated 18S rRNA gene sequences of Hepatocystis sp. from Tanzanian vervet monkeys were registered under the accession numbers OM281567 and OM281564 in GenBank. Although Hepatocystis infections do not cause clinical disease, they may interfere with the research data. The results of the current study pointed out that after proper nutrition and implementation of good physical environmental conditions for 3-4 months, the imported monkeys obviously gained weight and most of their hematological parameters, even in the presence of the parasite, returned to the normal levels and the experimental monkeys would be ready for use in studies., Competing Interests: The authors declare that they have no conflict of interest.

Serology and molecular tests are the two most commonly used methods for rapid COVID-19 infection testing. The two types of tests have different mechanisms to detect infection, by measuring the presence of viral SARS-CoV-2 RNA (molecular test) or detecting the presence of antibodies triggered by the SARS-CoV-2 virus (serology test). A handful of studies have shown that symptoms, combined with demographic and/or diagnosis features, can be helpful for the prediction of COVID-19 test outcomes. However, due to nature of the test, serology and molecular tests vary significantly. There is no existing study on the correlation between serology and molecular tests, and what type of symptoms are the key factors indicating the COVID-19 positive tests. In this study, we propose a machine learning based approach to study serology and molecular tests, and use features to predict test outcomes. A total of 2,467 donors, each tested using one or multiple types of COVID-19 tests, are collected as our testbed. By cross checking test types and results, we study correlation between serology and molecular tests. For test outcome prediction, we label 2,467 donors as positive or negative, by using their serology or molecular test results, and create symptom features to represent each donor for learning. Because COVID-19 produces a wide range of symptoms and the data collection process is essentially error prone, we group similar symptoms into bins. This decreases the feature space and sparsity. Using binned symptoms, combined with demographic features, we train five classification algorithms to predict COVID-19 test results. Experiments show that XGBoost achieves the best performance with 76.85% accuracy and 81.4% AUC scores, demonstrating that symptoms are indeed helpful for predicting COVID-19 test outcomes. Our study investigates the relationship between serology and molecular tests, identifies meaningful symptom features associated with COVID-19 infection, and also provides a way for rapid screening and cost effective detection of COVID-19 infection., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 Elsevier Inc. All rights reserved.)

Background: In the adjuvant setting of cancer treatment, de-escalation strategies have the goal of omitting or minimizing treatment in patients, without compromising outcomes. Historically, eligibility for adjuvant treatment solely relied on the patient's clinical and tumor's pathological characteristics. At the turn of the century, based on new biological understanding, molecular-based strategies were tested and sometimes implemented., Main Body: However, we illustrate how molecularly based de-escalation strategies may paradoxically lead to overtreatment. This may happen when the novel approach is tested in lieu of standard management and may not yield the same results when being implemented in addition to usual practice. In the DYNAMIC trial, adjuvant chemotherapy decision in stage II colon cancer was compared between a circulating tumor DNA (ctDNA)-based approach and the standard care. We show this may result in more patients receiving oxaliplatin-based chemotherapy and may expose a similar proportion of patients to chemotherapy if the novel strategy is implemented in addition to usual practice. The other potential risk is undertreatment. We provide an illustration of early breast cancer, where the decision of adjuvant chemotherapy based on the gene expression signature MammaPrint may lead to inferior outcomes as compared with the clinico-pathologic strategy. This may also happen when non-inferiority designs have large margins. Among solutions, it should be acknowledged that clinico-pathological features, like T4 in colon cancer, may not be abandoned and replaced by novel strategies in real-life practice. Therefore, novel strategies should be tested in addition to standard of care, and not in lieu of. Second, de-escalation trials should focus on the settings where the standard of care has a widespread agreement. This would avoid the risk of testing non-inferiority against an ineffective therapy, which guarantees successes without providing informative data., Conclusion: Simply because a molecular test is rational does not mean it can improve patient outcomes. Here, we highlight how molecular test-based strategies may result in either overtreatment or undertreatment. In the rapidly evolving field of medicine, where technological advances may be transformative, our piece highlights scientific pitfalls to be aware of when considering running such trials or before implementing novel strategies in daily practice., (© 2022. The Author(s).)

Bacterial vaginosis (BV) is the most common cause of abnormal vaginal discharge. BV represents a dysbiosis with the acquisition of a diverse community of anaerobic bacteria and a reduction in lactobacilli burden. Our objective was to evaluate the Aptima BV assay kit for the diagnosis of BV. From May to August 2019, we enrolled outpatients and inpatients, including nonpregnant women above 18 with vaginosis symptoms, consulting at Nantes University hospital. The Aptima BV assay measures the loads of Gardnerella vaginalis, Atopobium vaginae, and Lactobacillus species in relation to overall bacterial load. The Aptima BV assay was compared to Nugent scoring (NS). A total of 456 women were enrolled, and 347 patients met the inclusion criteria with data available for the analysis. NS was used to classify the samples and 144 (41.5%) samples were classified as normal (NS = 0-3), 45 (13%) as BV (NS = 7-10), 38 (11%) presented an intermediate vaginal microbiota (3 < NS < 7), 79 (22.7%) had various bacteria (excluding vaginal flora), 29 (8.3%) had insufficient bacterial density, and 12 (3.5%) had a predominance of yeasts. The Aptima BV kit displayed a sensitivity of 91.1% and specificity of 94.4% with a positive predictive value (PPV) of 83.7% and a negative predictive (NPV) value of 97.1%. The results of this monocentric retrospective study show that Aptima BV kit has a good diagnostic correlation compared to standard of care for dysbiotic diagnosis cases. IMPORTANCE The possibility exists of the involvement of a new molecular test in the routine algorithm of bacterial vaginosis diagnosis in microbiology laboratories. This manuscript reports on our experience, and we propose an organization combining Nugent scoring and molecular testing, especially for intermediate Nugent scores.

Rifampicin-resistant tuberculosis (RR-TB) has become a major threat globally. This study aims to develop a new assay, RIF-RDp, to enhance the detection of RR-TB based on combined locked nucleic acid (LNA) probes with high-resolution melting curve analysis (HRM). Two new LNA probes were designed to target the class-III and IV mutations of rpoB , H526D, and D516V. LNA probes showed 100% specificity in the detection of mutant targets among characterized and blinded Mycobacterium tuberculosis (Mtb) isolates. The performance of RIF-RDp was evaluated using 110 blinded clinical Mtb isolates in northern Thailand against drug-susceptibility testing (DST), DNA sequencing, and a commercial real-time PCR kit. This assay showed sensitivity and specificity of 94.55% and 98.18% compared to DST, and 96.36% and 100% compared to DNA sequencing. The efficacy of RIF-RDp was comparable to the commercial kit and DNA sequencing. The Cohen's Kappa statistic showed almost perfect agreement between RIF-RDp and the commercial kit (κ = 0.95), and RIF-RDp and DNA sequencing (κ = 0.96). Furthermore, this is the first report of the rare mutation profiles, S531W, and a triple codon deletion (510-512) in northern Thailand. According to high accuracy, the RIF-RDp assay may render an easy-to-use, low-cost, and promising diagnostics of RR-TB in the future.

Objective: Improved molecular testing for common somatic mutations and the identification of mRNA and microRNA expression classifiers are promising approaches for the diagnosis of thyroid nodules. However, there is a need to improve the diagnostic accuracy of such tests for identifying thyroid cancer. Recent findings have revealed a crucial role of long non-coding RNAs (lncRNAs) in gene modulation. Thus, we aimed to evaluate the diagnostic value of selected lncRNAs from The Atlas of Noncoding RNAs in Cancer (TANRIC) thyroid cancer dataset., Methods: LncRNAs in TANRIC thyroid cancer dataset that have significantly increased or decreased expression in papillary thyroid cancer (PTC) tissues were selected as candidates for PTC diagnosis. Surgical specimens from patients who underwent thyroidectomy were used to determine the separation capability of candidate lncRNAs between malignant and benign nodules. Fine needle aspiration samples were obtained and screened for candidate lncRNAs to verify their diagnostic value., Results: LRRC52-AS1, LINC02471, LINC02082, UNC5B-AS1, LINC02408, MPPED2-AS1, LNCNEF, LOC642484, ATP6V0E2-AS1, and LOC100129129 were selected as the candidate lncRNAs. LRRC52-AS1, LINC02082, UNC5B-AS1, MPPED2-AS1, LNCNEF, and LOC100129129 expression levels were significantly increased or decreased in malignant nodules compared to those in benign nodules and paired normal thyroid tissues. The combination of LRRC52-AS1, LINC02082, and UNC5B-AS1 showed favorable results for the diagnosis of PTC from fine needle aspirates, with 88.9% sensitivity and 100.0% specificity., Conclusions: LncRNA expression analysis is a promising approach for advancing the molecular diagnosis of PTC. Further studies are needed to identify lncRNAs of additional diagnostic value., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Kim, Yu, Kim, Hwang, Kim, Ku, Yoon, Kwak, Nam and Lee.)

Since the start of the COVID-19 pandemic, 10 manufacturers of molecular tests for SARS-CoV-2 have received Emergency Use Authorizations from the U.S. Food and Drug Administration for point-of-care or over the counter use. In this review, the working principle of these tests is described as well as the relevant characteristics (e.g. time-to-result and specimen type). The analytical (e.g. analytical sensitivity) and clinical performance (positive and negative percent agreement) and useability characteristics (e.g. cost, reusability and throughput) of these tests are compared and critically reviewed. Also details for relevant respiratory multiplex assays of these 10 manufacturers are discussed. Critical review of scientific literature on these authorized tests revealed that for many of these tests publications are almost or completely absent, with the exception of two systems. The Xpert Xpress has been thoroughly investigated and good performance has been reported, whereas ID NOW is also well-represented in literature, but has relatively low sensitivity., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 The Author(s).)

Background: Until 2016, microscopy was the main tool for the early detection of pulmonary tuberculosis in Cameroon, especially in remote settings. Due to the poor sensitivity of microscopy, there was a need to implement a molecular assay in order to improve tuberculosis case detection., Intervention: In 2017, tuberculosis loop-mediated isothermal amplification (TB-LAMP), a molecular rapid diagnostic test recommended by the World Health Organization, was implemented in Cameroon as a replacement test of microscopy for initial diagnosis of pulmonary tuberculosis and also as a follow-on test to microscopy for smear-negative sputum specimens. A roll out plan for TB-LAMP implementation in Cameroon had been developed from January 2017 to April 2017, followed by initial implementation at four sites in May 2017. Additional sites were added progressively., Lessons Learnt: The use of TB-LAMP as a follow-on test to microscopy for smear-negative sputum specimens helped in the detection of tuberculosis in 14.77% of those who were sputum-smear negative in 2019. Tuberculosis-loop-mediated isothermal amplification usage as an initial test, followed by testing with Xpert MTB/RIF for rapid tuberculosis and rifampicin resistance detection during tuberculosis mass screening campaigns, reduced the turn-around time by 73.23% as compared to when the Gene Xpert instrument was used alone., Recommendations: The implementation and scaling up of TB-LAMP in Cameroon contributed to increase access to tuberculosis molecular diagnosis in remote settings and as such improved tuberculosis case notification. However, to better enhance this notification and optimise the use of a TB-LAMP instrument, a suitable sample transport system is recommended., Competing Interests: The authors declare that they have no financial or personal relationships that may have inappropriately influenced them in writing this article., (© 2022. The Authors.)

Introduction: Rapid molecular tests could accelerate the control of extended-spectrum beta-lactamase-producing Enterobacterales (ESBL-PE) and carbapenemase-producing organisms (CPO) in intensive care units (ICUs)., Objective and Methods: This interventional 12-month cohort study compared a loop-mediated isothermal amplification (LAMP) assay performed directly on rectal swabs with culturing methods (control period, 6 months), during routine ICU screening. Contact precautions (CP) were implemented for CPO or non-E. coli ESBL-producing Enterobacterales (nEcESBL-PE) carriers. Using survival analysis, we compared the time intervals from admission to discontinuation of unnecessary preemptive CP among patients at-risk and the time intervals from screening to implementation of CP among newly identified carriers. We also compared diagnostic performances, and nEcESBL-PE/CPO acquisition rates. This study is registered, ISRCTN 23588440., Results: We included 1043 patients. During the intervention and control phases, 92/147 (62.6%) and 47/86 (54.7%) of patients at-risk screened at admission were candidates for early discontinuation of preemptive CP. The LAMP assay had a positive predictive value (PPV) of 44.0% and a negative predictive value (NPV) of 99.9% for CPO, and 55.6% PPV and 98.2% NPV for nEcESBL-PE. Due to result notification and interpretation challenges, the median time from admission to discontinuation of preemptive CP increased during the interventional period from 80.5 (95% CI 71.5-132.1) to 88.3 (95% CI 57.7-103.7) hours (p = 0.47). Due to the poor PPV, we had to stop using the LAMP assay to implement CP. No difference was observed regarding the incidence of nEcESBL-PE and CPO acquisition., Conclusion: A rapid screening strategy with LAMP assays performed directly on rectal swabs had no benefit for infection control in a low-endemicity setting., (© 2022. The Author(s).)

Pregnancy-associated plasma protein A (PAPPA) acts as an oncogene, and its expression is increased in multiple malignancies, including thyroid cancer. Molecular tests represent a useful tool in the management of indeterminate thyroid nodules; however, they are not conducted in all centers, and they contribute to increase the per-patient cost of nodule evaluation. In this study, we examined whether PAPPA expression could represent a promising new screening test in the management of indeterminate thyroid nodules. Toward this aim, PAPPA expression was evaluated in 107 fine needle aspiration cytologies (FNAC) belonging to Bethesda III-IV categories that had been sent to molecular biology to discriminate the nature of the nodules. We found that the PAPPA expression increased and showed an elevated sensitivity (97.14%) and negative predictive value (98%) in indeterminate cytological samples positive for mutations. The enhanced expression was not linked to a specific oncogene. Our findings demonstrated that assessing the PAPPA expression in indeterminate thyroid cytologies could represent a useful screening tool to select all patients that effectively need to be sent to molecular testing, thereby, leading to a potential cost reduction in the management of patients.

Diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) during border screening among returning residents and prioritized travelers during the early phase of a pandemic can reduce the risk of importation and transmission in the community. This study aimed to compare the accuracy of various SARS-CoV-2 diagnostics and assess their potential utility as border screening for infection and immunity. Systematic literature searches were conducted in six electronic databases for studies reporting SARS-CoV-2 diagnostics (up to April 30, 2020). Meta-analysis and methodological assessment were conducted for all included studies. The performance of the diagnostic tests was evaluated with pooled sensitivity, specificity, and their respective 95% confidence intervals. A total of 5,416 unique studies were identified and 95 studies (at least 29,785 patients/samples) were included. Nucleic acid amplification tests (NAAT) consistently outperformed all other diagnostic methods regardless of the selected viral genes with a pooled sensitivity of 98% and a pooled specificity of 99%. Point-of-care (POC) serology tests had moderately high pooled sensitivity (69%), albeit lower than laboratory-based serology tests (89%), but both had high pooled specificity (96-98%). Serology tests were more sensitive for sampling collected at ≥ 7 days than ≤ 7 days from the disease symptoms onset. POC NAAT and POC serology tests are suitable for detecting infection and immunity against the virus, respectively as border screening. Independent validation in each country is highly encouraged with the preferred choice of diagnostic tool/s., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Chua, Gwee, Wang, Gui and Pang.)

Background: Asymptomatic Leishmania infection may play an important role in the transmission of the parasite in endemic areas. At present there is no consensus on the definition of asymptomatic Leishmania infection, nor is there a safe and accessible gold standard test for its identification., Methods: This paper presents a scoping review to summarize definitions of asymptomatic Leishmania infection found in the literature, as well as to detail the approach (molecular, serological, cellular, and/or parasitological tests) used by researchers to identify this asymptomatic population. A scoping review of published and gray literature related to asymptomatic Leishmania infection was conducted; retrieved citations were screened based on predefined eligibility criteria, and relevant data items were extracted from eligible articles. The analysis is descriptive and is presented using tables, figures, and thematic narrative synthesis., Results: We conducted a screening of 3008 articles, of which 175 were selected for the full review. Of these articles, we selected 106 that met the inclusion criteria. These articles were published between 1991 and 2021, and in the last 5 years, up to 38 articles were reported. Most of the studies were conducted in Brazil (26%), Spain (14%), India (12%), Bangladesh (10%), and Ethiopia (7%). Of the studies, 84.9% were conducted in the immunocompetent population, while 15.1% were conducted in the immunosuppressed population (HIV, immunosuppressive drugs, and organ transplantation population). We report 14 different techniques and 10 strategies employed by researchers to define asymptomatic Leishmania infection in an endemic area., Conclusions: The definition of asymptomatic Leishmania infection is not unified across the literature, but often includes the following criteria: residence (or extended stay) in a Leishmania-endemic area, no reported signs/symptoms compatible with leishmaniasis, and positive on a combination of serological, molecular, cellular, and/or parasitological tests. Caution is recommended when comparing results of different studies on the subject of asymptomatic infections, as the reported prevalence cannot be confidently compared between areas due to the wide variety of tests employed by research groups. More research on the importance of asymptomatic immunosuppressed and immunocompetent Leishmania-positive populations in leishmaniasis epidemiology is required., (© 2021. The Author(s).)

The increased demand for SARS-CoV-2 molecular testing during the COVID-19 pandemic resulted in shortage of reagents and consumables. Pooling of specimens could be an alternative strategy to overcome these problems. Initial evaluation of the pooling strategy was performed using known positive specimens, previously tested individually, and their respective pools of plus four (5X), five (6X) and nine (10X) known negative specimens. Subsequently, 35 positive 5X and 35 positive 6X pools containing only one positive specimen per pool were analyzed prospectively regarding the difference in Ct values in pooled versus individual specimens. When the number of samples in the pool were five or six, the average deviation of Ct differences was < 1; therefore, this strategy was followed in the prospective study. Significant difference in Ct values was observed in positive specimens when tested individually and in 5X pools ( p  = 0.006), while the difference was not significant when positive specimens were tested individually and in 6X pools ( p  = 0.07). The difference in Ct values was not significant between the 5X and 6X pools. Testing in pools of five or six specimens is a reliable option for SARS-CoV-2 RNA detection when mass testing is needed., Competing Interests: Conflict of interestThe authors declare that there is no conflict of interest., (© Indian Virological Society 2021.)

The coronavirus disease 2019 (COVID-19) pandemic unfolded due to the widespread severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission reinforced the urgent need for affordable molecular diagnostic alternative methods for massive testing screening. We present the clinical validation of a pH-dependent colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) for SARS-CoV-2 detection. The method revealed a limit of detection of 19.3 ± 2.7 viral genomic copies/μL when using RNA extracted samples obtained from nasopharyngeal swabs collected in guanidine-containing viral transport medium. Typical RT-LAMP reactions were performed at 65°C for 30 min. When compared to reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR), up to cycle-threshold (Ct) value 32, RT-LAMP presented 98% [95% confidence interval (CI) = 95.3-99.5%] sensitivity and 100% (95% CI = 94.5-100%) specificity for SARS-CoV-2 RNA detection targeting E and N genes. No cross-reactivity was detected when testing other non-SARS-CoV virus, confirming high specificity. The test is compatible with primary RNA extraction-free samples. We also demonstrated that colorimetric RT-LAMP can detect SARS-CoV-2 variants of concern and variants of interest, such as variants occurring in Brazil named gamma (P.1), zeta (P.2), delta (B.1.617.2), B.1.1.374, and B.1.1.371. The method meets point-of-care requirements and can be deployed in the field for high-throughput COVID-19 testing campaigns, especially in countries where COVID-19 testing efforts are far from ideal to tackle the pandemics. Although RT-qPCR is considered the gold standard for SARS-CoV-2 RNA detection, it requires expensive equipment, infrastructure, and highly trained personnel. In contrast, RT-LAMP emerges as an affordable, inexpensive, and simple alternative for SARS-CoV-2 molecular detection that can be applied to massive COVID-19 testing campaigns and save lives., Competing Interests: HM is part of Visuri company. Results presented here are the basis of a COVID-19 RT-LAMP diagnostic test offered by Visuri named OmniLAMP® SARS-CoV-2 kit. PA and RM-N are co-founders and scientific advisors at CEPHA Biotech. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Alves, de Oliveira, Franco-Luiz, Almeida, Gonçalves, Borges, Rocha, Rocha, Bezerra, Miranda, Capanema, Martins, Weber, Teixeira, Wallau and do Monte-Neto.)

Group A streptococcus (GAS) causes significant morbidity and mortality in New Zealand and is responsible for invasive disease and immune sequelae, including acute rheumatic fever (ARF). Early treatment of GAS pharyngitis reduces the risk of ARF. In settings with a high burden of GAS disease, a rapid GAS pharyngitis diagnostic test with a strong negative predictive value is needed to enable prompt and accurate treatment. This prospective study compares the Xpert Xpress Strep A molecular test (Cepheid) to throat culture and a second molecular method, the BioGX group A streptococcus-open system reagent (OSR) for BD Max for the diagnosis of GAS pharyngitis. Throat swabs were collected from the emergency department and wards of Middlemore Hospital, New Zealand. The BioGX group A streptococcus OSR for BD Max contributes to the composite gold standard of throat culture or both molecular methods positive. Basic demographic, clinical, and laboratory data were collected. Two hundred five out of two hundred fourteen swabs were suitable for analysis. Of those, 28/205 (13.7%) were GAS culture positive, 45/205 (22%) Xpert Xpress Strep A positive, and 38/205 (18.5%) BioGX positive. Compared to culture, the sensitivity, specificity, and positive and negative predictive values of the Xpert Xpress Strep A molecular test were 100%, 90.4%, 62.2%, and 100%, respectively. Compared to the composite gold standard, the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were 100%, 95.8%, 84.4%, and 100%, respectively. Seventeen samples were Xpert Xpress positive but culture negative; 6 of these 17 swabs represent true positives with evidence of recent GAS infection. Ten samples were culture negative but both Xpert Xpress and BioGX positive. The Xpert Xpress Strep A molecular test is highly sensitive with a strong negative predictive value and rapid turnaround time. It can be safely introduced as a first-line test for throat swabs in a high-incidence ARF population.