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PURPOSE: Unresectable pediatric low-grade gliomas (LGG) usually need adjuvant therapy, and carboplatin hypersensitivity reaction (HR) commonly leads to premature treatment cessation of a standard chemotherapy regimen. In the molecular era, advances in understanding tumor genetic characteristics allowed the development of targeted therapies for this group of tumors; however, cost-effectiveness assessment of treatments, especially in low-income countries, is crucial. The aim is to describe the results of carboplatin desensitization protocol in a single center in a middle-income country. METHOD: Prospective analysis of children with LGG submitted to carboplatin desensitization from December 2017 to June 2020 with follow-up until April 2024. RESULTS: Nine patients were included. The mean age was 11 years. Five patients were male. Seven had optic pathway and two cervicomedullary location. Six had histologic diagnosis and four molecular analyses. The incidence of carboplatin reactions during the study period was 39.1%. Six patients underwent skin prick test, three with positive results. The first HR occurred, on average, around the 9th cycle of treatment. All patients had cutaneous symptoms, and five out of nine had anaphylaxis as the first reaction. 77.7% of the patients completed the protocol, and the clinical benefit rate (stable disease and partial response) was 88.8%. Six patients further required other lines of therapy. Monthly, the total cost for carboplatin was $409.09, and for target therapies (dabrafenib plus trametinib), $4929.28 to $5548.57. CONCLUSION: Our study presented an interesting and cost-effective option where desensitization allowed children with HR to be treated with first-line therapy, avoiding the discontinuation of an effective treatment.

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Fusarium and Neocosmospora are two fungal genera recently recognized in the list of fungal priority pathogens. They cause a wide range of diseases that affect humans, animals, and plants. In clinical laboratories, there is increasing concern about diagnosis due to limitations in sample collection and morphological identification. Despite the advances in molecular diagnosis, due to the cost, some countries cannot implement these methodologies. However, recent changes in taxonomy and intrinsic resistance to antifungals reveal the necessity of accurate species-level identification. In this review, we discuss the current phenotypic and molecular tools available for diagnosis in clinical laboratory settings and their advantages and disadvantages.

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Fasciola hepatica has a complex lifecycle with multiple intermediate and definitive hosts and influenced by environmental factors. The disease causes significant morbidity in children and its prevalent worldwide. There is lack of data about distribution and burden of the disease in endemic regions, owing to poor efficacy of the different diagnostic methods used. A novel PCR-based test was developed by using a portable mini-PCR® platform to detect Fasciola sp. DNA and interpret the results via a fluorescence viewer and smartphone image analyzer application. Human stool, snail tissue, and water samples were used to extract DNA. Primers targeting the ITS-1 of the 18S rDNA gene of Fasciola sp. were used. The limit of detection of the mini-PCR test was 1 fg/µL for DNA samples diluted in water, 10 fg/µL for Fasciola/snail DNA scramble, and 100 fg/µL for Fasciola/stool DNA scramble. The product detection by agarose gel, direct visualization, and image analyses showed the same sensitivity. The Fh mini-PCR had a sensitivity and specificity equivalent to real-time PCR using the same specimens. Testing was also done on infected human stool and snail tissue successfully. These experiments demonstrated that Fh mini-PCR is as sensitive and specific as real time PCR but without the use of expensive equipment and laboratory facilities. Further testing of multiple specimens with natural infection will provide evidence for feasibility of deployment to resource constrained laboratories.

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BACKGROUND: Diagnosing imprinting defects in neonates and young children presents challenges, often necessitating molecular analysis for a conclusive diagnosis. The isolation of genetic material from oral swabs becomes crucial, especially in settings where blood sample collection is impractical or for vulnerable populations like newborns, who possess limited blood volumes and are often too fragile for invasive procedures. Oral swab samples emerge as an excellent source of DNA, effectively overcoming obstacles associated with rare diseases. METHODS: In our study, we specifically addressed the determination of the quality and quantity of DNA extracted from oral swab samples using NaCl procedures. RESULTS: We compared these results with extractions performed using a commercial kit. Subsequently, the obtained material underwent MS-HRM analysis for loci associated with imprinting diseases such as Prader-Willi and Angelman syndromes. CONCLUSIONS: Our study emphasizes the significance of oral swab samples as a reliable source for obtaining DNA for MS-HRM analysis. NaCl extraction stands out as a practical and cost-effective method for genetic studies, contributing to a molecular diagnosis that proves particularly beneficial for patients facing delays in characterization, ultimately influencing their treatment.

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BACKROUND: Stored product protection from insect pests relies heavily on the use of phosphine. The most serious drawback of phosphine is the development of resistance in major stored product insects worldwide, including the red flour beetle, Tribolium castaneum (Herbst) and the lesser grain borer, Rhyzopertha dominica (F.). Two genetic loci are responsible for phosphine resistance: the rph1 (S349G mutation in the cyt-b5-r homolog) in T. castaneum and the rph2 (P45/49S mutation in the dihydrolipoamide dehydrogenase (dld) gene) in T. castaneum and R. dominica. RESULTS: In this study, we have developed and applied high-throughput, practical and specific molecular diagnostics (TaqMan qPCR) for monitoring mutations S349G, P45S and P49S. In our pilot monitoring application, we have included phosphine-resistant and susceptible populations from different parts of the world (USA, Australia, Brazil) and European strains from Greece and Serbia. Our results for the resistant T. castaneum showed a P45S mutant allele frequency (MAF) of 100% and 75.0% in the populations from Serbia and Brazil, respectively. Regarding the susceptible T. castaneum, P45S was detected in Greece (MAF = 62.5%) and was absent in Australia (MAF = 0.0%). Additionally, the S349G mutation was found to be fixed in all resistant populations, while it was also detected in susceptible ones (frequencies: 65.0% and 100.0%). The only case where both mutations were fixed (100%) was a resistant population from Serbia. In R. dominica, the P49S mutation was found only in the two resistant R. dominica populations from Serbia and Greece (50.0% and 100%) and was absent from the susceptible one from Greece; thus, P49S seems to be a satisfactory indicator for monitoring phosphine resistance. CONCLUSIONS: Our P49S detection assay in R. dominica seems to be a viable option in this direction, yet its utilization needs additional large-scale confirmatory work. The identification of additional resistance markers also should be prioritized. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

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In the evolution of modern medicine, artificial intelligence (AI) has been proven to provide an integral aspect of revolutionizing clinical diagnosis, drug discovery, and patient care. With the potential to scrutinize colossal amounts of medical data, radiological and histological images, and genomic data in healthcare institutions, AI-powered systems can recognize, determine, and associate patterns and provide impactful insights that would be strenuous and challenging for clinicians to detect during their daily clinical practice. The outcome of AI-mediated search offers more accurate, personalized patient diagnoses, guides in research for new drug therapies, and provides a more effective multidisciplinary treatment plan that can be implemented for patients with chronic diseases. Among the many promising applications of AI in modern medicine, medical imaging stands out distinctly as an area with tremendous potential. AI-powered algorithms can now accurately and sensitively identify cancer cells and other lesions in medical images with greater accuracy and sensitivity. This allows for earlier diagnosis and treatment, which can significantly impact patient outcomes. This review provides a comprehensive insight into diagnostic, therapeutic, and ethical issues with the advent of AI in modern medicine.

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In the absence of validated biomarkers to control the cure of Chagas disease, PCR-based diagnosis is being used as the main tool for an early indication of therapeutic failure. However, since it is considered a technique of complex reproducibility, mainly due to difficulties in establishing accurate controls to guarantee the quality of the reaction, the use of PCR for Chagas disease diagnosis is restricted to specialized centers. In an effort to disseminate the molecular diagnosis of Chagas disease and its applications, new diagnostic kits based on qPCR have been made available in the market in recent years. Here, we show the results of the validation of the NAT Chagas kit (Nucleic Acid Test for Chagas Disease) for the detection and quantification of T. cruzi in blood samples of patients suspected of Chagas disease infection. The kit, composed of a TaqMan duplex reaction targeting the T. cruzi satellite nuclear DNA and an exogenous internal amplification control, presented a reportable range from 104 to 0.5 parasite equivalents/mL and a limit of detection (LOD) of 0.16 parasite equivalents/mL of blood. In addition, the NAT Chagas kit detected T. cruzi belonging to all six discrete typing units (DTUs-TcI to TcVI), similarly to the in-house real-time PCR performed with commercial reagents, which has been selected as the best performance assay in the international consensus for the validation of qPCR for Chagas disease. In the clinical validation presented here, the kit showed 100% sensitivity and 100% specificity when compared to the consensus in-house real-time PCR assay. Thus, the NAT Chagas kit, which is produced entirely in Brazil under the international standards of good manufacturing practices (GMP), appears as an excellent alternative to enable the molecular diagnosis of Chagas disease in public and private diagnostic centers, as well as to facilitate the monitoring of patients under etiological treatment participating in clinical trials.

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In resource-limited conditions such as the COVID-19 pandemic, on-site detection of diseases using the Point-of-care testing (POCT) technique is becoming a key factor in overcoming crises and saving lives. For practical POCT in the field, affordable, sensitive, and rapid medical testing should be performed on simple and portable platforms, instead of laboratory facilities. In this review, we introduce recent approaches to the detection of respiratory virus targets, analysis trends, and prospects. Respiratory viruses occur everywhere and are one of the most common and widely spreading infectious diseases in the human global society. Seasonal influenza, avian influenza, coronavirus, and COVID-19 are examples of such diseases. On-site detection and POCT for respiratory viruses are state-of-the-art technologies in this field and are commercially valuable global healthcare topics. Cutting-edge POCT techniques have focused on the detection of respiratory viruses for early diagnosis, prevention, and monitoring to protect against the spread of COVID-19. In particular, we highlight the application of sensing techniques to each platform to reveal the challenges of the development stage. Recent POCT approaches have been summarized in terms of principle, sensitivity, analysis time, and convenience for field applications. Based on the analysis of current states, we also suggest the remaining challenges and prospects for the use of the POCT technique for respiratory virus detection to improve our protection ability and prevent the next pandemic.

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Periprosthetic joint infection (PJI) remains one of the most common complications of total knee arthroplasty. Although mainly caused by Staphylococcus aureus and other Gram-positive microorganisms, occasionally, commensal or environmental bacteria are reported as causative agents of these infections. The present work aimed to report a case of PJI caused by an imipenem-resistant Mycobacterium senegalense strain. A bacterial strain isolated from the culture of intraoperative samples was observed by optical microscopy after Gram and Ziehl-Neelsen staining. The species identification was performed by mass spectrometry analysis and partial sequencing of the heat shock protein 65 (hsp65) gene. The antimicrobial profile of the clinical isolate was determined according to the Clinical and Laboratory Standards Institute. Mass spectrometry and gene sequencing analysis identified the bacterial isolate as Mycobacterium fortuitum complex and M. senegalense, respectively. The isolated was found exhibiting an imipenem-resistant profile. The accurate and timely identification, as well as investigation of the antimicrobial susceptibility profile, of fast-growing nontuberculous mycobacteria species are crucial for establishing the prompt and correct treatment of the infection, particularly in cases of patients at greater risk for opportunistic and severe infections.

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Introducción. Las infecciones de transmisión sexual (ITS) son y seguirán siendo un serio problema de salud pública en todo el mundo según los datos de la OMS, con el agravante que la mayoría de los casos son asintomáticos y, además, no existe otro reservorio distinto al humano. El diagnóstico se puede realizar con pruebas tradicionales y moleculares, estas últimas incluyen la reacción en cadena de la polimerasa (PCR), de las cuales existen varios tipos, entre ellas, la PCR múltiple que tiene la capacidad de detectar ITS polimicrobianas a partir de una sola muestra. El objetivo de este estudio fue establecer cuáles fueron las infecciones de transmisión sexual más frecuentes en diferentes grupos de pacientes, así como determinar la utilidad del uso de la técnica de PCR múltiple en el diagnóstico de las ITS. Metodología. Se trata de un estudio observacional de corte transversal realizado entre los años 2021 y 2022 con pacientes que acudieron al servicio de diagnóstico del Laboratorio Clínico VID por sospecha de ITS. Las muestras recolectadas fueron evaluadas utilizando una prueba comercial basada en la técnica de PCR múltiple e hibridación. Las muestras procesadas fueron: orina e hisopados de endocérvix, uretra, recto, faringe y úlceras. Resultados. Se estudiaron 1.027 pacientes, de estos, 228 (22,2 %) fueron positivos para diferentes agentes de trasmisión sexual, distribuidos así: 50 (21,9 %) mujeres, 129 (56,6 %) hombres heterosexuales y 49 (21,5 %) hombres que tenían sexo con hombres (HSH). La edad promedio de las mujeres fue 30 años, y la de ambos grupos de hombres fue 36 años. Los microorganismos más frecuentemente identificados en mujeres fueron: C. trachomatis (A-K) en 28,6 %, seguido de virus herpes simplex tipo 2 (VHS-2) en 26,8 % y N. gonorrhoeae en 17,9 %. En hombres heterosexuales fueron C. trachomatis (A-K) en 37,5 %, N. gonorrhoeae en 21,5 % y VHS-2 en 18,7 %. En HSH fueron C. trachomatis (L1-L3) en 32,7 %, seguido de N. gonorrhoeae en 27,6 %, y de C. trachomatis (A-K) y VHS-2, ambos en 13,8 %. En 11 hombres heterosexuales, 8 HSH y en 6 mujeres, se identificó infección polimicrobiana. Conclusiones. C. trachomatis (A-K) fue el microorganismo más prevalente causante de ITS, seguido de N. gonorrhoeae en ambos grupos de hombres, y de VHS-2 en las mujeres, muy similar a lo reportado a nivel mundial. La prueba de PCR múltiple permite la detección de infecciones polimicrobianas comúnmente asociadas a ITS y el diagnóstico es preciso y confiable, incluso en pacientes asintomáticos

Sexually transmitted infections (STIs) are and will continue to be a serious public health problem throughout the world according to WHO data, with the aggravating factor that most cases are asymptomatic and, furthermore, there is no other reservoir other than humans. The diagnosis can be made with traditional and molecular tests, the latter include the polymerase chain reaction (PCR), of which there are several types, among them, multiplex PCR that has the capacity to detect polymicrobial STIs from a single sample. The objective of this study was to establish which were the most frequent sexually transmitted infections in different groups of patients, as well as to determine the usefulness of the multiplex PCR technique in the diagnosis of STIs. Methodology. This is an observational, cross-sectional study carried out between 2021 and 2022 with patients who attended the VID Clinical Laboratory for suspected STIs. The collected samples were evaluated using a commercial test based on the multiplex PCR technique and hybridization. The samples processed were: urine and swabs from endocervix, urethra, rectum, pharynx, and ulcers. Results. The study included 1,027 patients, of these, 228 (22.2%) were positive for different sexually transmitted agents, distributed as follows: 50 (21.9%) women, 129 (56.6%) heterosexual men and 49 (21.5%) men who had sex with men (MSM). The average age of the women was 30 years, and that of both groups of men was 36 years. The microorganisms most frequently identified in women were: C. trachomatis (A-K) in 28.6%, followed by herpes simplex virus type 2 (HSV-2) in 26.8% and N. gonorrhoeae in 17.9%. In heterosexual men they were C. trachomatis (A-K) in 37.5%, N. gonorrhoeae in 21.5% and HSV-2 in 18.7%. In MSM they were C. trachomatis (L1-L3) in 32.7%, followed by N. gonorrhoeae in 27.6%, and C. trachomatis (A-K) and HSV-2, both in 13.8%. Polymicrobial infection was identified in 11 heterosexual men, 8 MSM, and 6 women. Conclusions. C. trachomatis (A-K) was the most prevalent STI-causing microorganism, followed by N. gonorrhoeae in both groups of men, and HSV-2 in women, very similar to that reported worldwide. The multiplex PCR test allows the detection of polymicrobial infections commonly associated with STIs and the diagnosis is accurate and reliable, even in asymptomatic patients

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Background: Accurate diagnosis followed by timely treatment is an effective strategy for the prevention of complications together with reducing schistosomiasis transmission. Recombinase Polymerase Amplification (RPA) is a simple, rapid, sensitive, and specific isothermal method with low resource needs. This research aimed at the development and optimisation of a real-time (RT) and a lateral flow (LF) RPA assay for the detection of Schistosoma mansoni. Methodology: Recombinase Polymerase Amplification reactions were performed at full- (as recommended) and half-volumes (to reduce costs), with RT or LF detection systems targeting the S. mansoni mitochondrial minisatellite region. The specificity was assessed using gDNA from other Schistosoma species, helminths co-endemic with S. mansoni, human stool, and urine, and Biomphalaria snail hosts. The analytical sensitivity was evaluated using serial dilutions of gDNA, synthetic copies of the target, and single eggs. The ability of both assays to detect the S. mansoni DNA in human urine and stool samples was also tested. The long-term stability of the RT-RPA reagents was evaluated by storing the reaction components in different temperature conditions for up to 3 weeks. Results: The RT- and the LF-RPA (SmMIT- and SmMIT-LF-RPA, respectively) presented similar results when used full- and half-volumes, thus the latter was followed in all experiments. The SmMIT-RPA was 100% specific to S. mansoni, able to detect a single egg, with a limit of detection (LOD) of down to 1 fg of gDNA and one synthetic copy of the target. The assay was able to detect S. mansoni DNA from stool containing 1 egg/g and in spiked urine at a concentration of 10 fg/µl. SmMIT-RPA reagents were stable for up to 3 weeks when kept at 19°C, and 2 weeks when stored at 27°C. The SmMIT-LF-RPA cross-reacted with Clinostomidae, presented the LOD of 10 fg and one synthetic copy of the target, being able to detect a single egg and 1 egg/g in a stool sample. The LOD in spiked urine samples was 10 pg/µl. Conclusion: The half-volume SmMIT-RPA is a promising method to be used in the field. It is specific, sensitive, robust, and tolerant to inhibitors, with a long-term stability of the reaction components and the real-time visualisation of results.

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Tegumentary leishmaniasis, a disease caused by protozoan parasites of the genus Leishmania, is a major public health problem in many regions of Latin America. Its diagnosis is difficult given other conditions resembling leishmaniasis lesions and co-occurring in the same endemic areas. A combination of parasitological and molecular methods leads to accurate diagnosis, with the latter being traditionally performed in centralized reference and research laboratories as they require specialized infrastructure and operators. Clustered regularly interspaced short palindromic repeats/CRISPR-associated protein (CRISPR/Cas) systems have recently driven innovative tools for nucleic acid detection that combine high specificity, sensitivity and speed and are readily adaptable for point-of-care testing. Here, we harnessed the CRISPR-Cas12a system for molecular detection of Leishmania spp., emphasizing medically relevant parasite species circulating in Peru and other endemic areas in Latin America, with Leishmania (Viannia) braziliensis being the main etiologic agent of cutaneous and mucosal leishmaniasis. We developed two assays targeting multi-copy targets commonly used in the molecular diagnosis of leishmaniasis: the 18S ribosomal RNA gene (18S rDNA), highly conserved across Leishmania species, and a region of kinetoplast DNA (kDNA) minicircles conserved in the L. (Viannia) subgenus. Our CRISPR-based assays were capable of detecting down to 5 × 10-2 (kDNA) or 5 × 100 (18S rDNA) parasite genome equivalents/reaction with PCR preamplification. The 18S PCR/CRISPR assay achieved pan-Leishmania detection, whereas the kDNA PCR/CRISPR assay was specific for L. (Viannia) detection. No cross-reaction was observed with Trypanosoma cruzi strain Y or human DNA. We evaluated the performance of the assays using 49 clinical samples compared to a kDNA real-time PCR assay as the reference test. The kDNA PCR/CRISPR assay performed equally well as the reference test, with positive and negative percent agreement of 100%. The 18S PCR/CRISPR assay had high positive and negative percent agreement of 82.1% and 100%, respectively. The findings support the potential applicability of the newly developed CRISPR-based molecular tools for first-line diagnosis of Leishmania infections at the genus and L. (Viannia) subgenus levels.

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Coronavirus disease (COVID-19) is an infectious disease caused by SARS-CoV-2. In Colombia, many commercial methods are now available to perform the RT-qPCR assays, and laboratories must evaluate their diagnostic accuracy to ensure reliable results for patients suspected of being positive for COVID-19. The purpose of this study was to compare four commercial RT-qPCR assays with respect to their ability to detect the SARS-CoV2 virus from nasopharyngeal swab samples referred to Laboratorio Carvajal IPS, SAS in Tunja, Boyacá, Colombia. We utilized 152 respiratory tract samples (Nasopharyngeal Swabs) from patients suspected of having SARS-CoV-2. The diagnostic accuracy of GeneFinderTM COVID-19 Plus RealAmp (In Vitro Diagnostics) (GF-TM), One-Step Real-Time RT-PCR (Vitro Master Diagnostica) (O-S RT-qPCR), and the Berlin modified protocol (BM) were assessed using the gold-standard Berlin protocol (Berlin Charité Probe One-Step RT-qPCR Kit, New England Biolabs) (BR) as a reference. Operational characteristics were estimated in terms of sensitivity, specificity, agreement, and predictive values. Using the gold-standard BR as a reference, the sensitivity/specificity of the diagnostic tests was found to be 100%/92.7% for GF-TM, 92.75%/67.47% for O-S RT-qPCR, and 100%/96.39% for the BM protocol. Using BR as a reference, the sensitivity/specificity for the diagnostic tests were found to be 100%/92.7% for the GF-TM assay, 92.72%/67.47% for the O-S RT-qPCR, and 100%/96.39% for BM. Relative to the BR reference protocol, the GF-TM and BM RT-PCR assays obtained similar results (k = 0.92 and k = 0.96, respectively), whereas the results obtained by O-S-RT-qPCR were only moderately similar. We conclude that the GF-TM and BM protocols offer the best sensitivity and specificity, with similar results in comparison to the gold-standard BR protocol. We recommend evaluating the diagnostic accuracy of the OS-RT-qPCR protocol in future studies with a larger number of samples.

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Sporotrichosis is the most important subcutaneous mycosis that affects humans and animals worldwide. The mycosis is caused after a traumatic inoculation of fungal propagules into the host and may follow an animal or environmental transmission route. The main culprits of sporotrichosis are thermodimorphic Sporothrix species embedded in a clinical clade, including S. brasiliensis, S. schenckii, S. globosa, and S. luriei. Although sporotrichosis occurs worldwide, the etiological agents are not evenly distributed, as exemplified by ongoing outbreaks in Brazil and China, caused by S. brasiliensis and S. globosa, respectively. The gold standard for diagnosing sporotrichosis has been the isolation of the fungus in vitro. However, with the advance in molecular techniques, molecular assays have complemented and gradually replaced the classical mycological tests to quickly and accurately detect and/or differentiate molecular siblings in Sporothrix. Nearly all techniques available for molecular diagnosis of sporotrichosis involve PCR amplification, which is currently moving towards detecting Sporothrix DNA directly from clinical samples in multiplex qPCR assays. From an epidemiological perspective, genotyping is key to tracing back sources of Sporothrix infections, detecting diversity in outbreak areas, and thus uncovering finer-scale epidemiological patterns. Over the past decades, molecular epidemiological studies have provided essential information to policymakers regarding outbreak management. From high-to-low throughput genotyping methods, MLSA, AFLP, SSR, RAPD, PCR-RFLP, and WGS are available to assess the transmission dynamics and sporotrichosis expansion. This review discusses the trends in the molecular diagnosis of sporotrichosis, genotyping techniques applied in molecular epidemiological studies, and perspectives for the near future.

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BACKGROUND: The diagnosis of cancer in Bethesda III/IV thyroid nodules is challenging as fine-needle aspiration (FNA) has limitations, and these cases usually require diagnostic surgery. As approximately 77% of these nodules are not malignant, a diagnostic test accurately identifying benign thyroid nodules can reduce "potentially unnecessary" surgery rates. We have previously reported the development and validation of a microRNA-based thyroid classifier (mir-THYpe) with high sensitivity and specificity, which could be performed directly from FNA smear slides. We sought to evaluate the performance of this test in real-world clinical routine to support clinical decisions and to reduce surgery rates. METHODS: We designed a real-world, prospective, multicentre study. Molecular tests were performed with FNA samples prepared at 128 cytopathology laboratories. Patients were followed-up from March 2018 until surgery or until March 2020 (patients with no indication for surgery). The final diagnosis of thyroid tissue samples was retrieved from postsurgical anatomopathological reports. FINDINGS: A total of 435 patients (440 nodules) classified as Bethesda III/IV were followed-up. The rate of avoided surgeries was 52·5% for all surgeries and 74·6% for "potentially unnecessary" surgeries. The test achieved 89·3% sensitivity, 81·65% specificity, 66·2% positive predictive value, and 95% negative predictive value. The test supported 92·3% of clinical decisions. INTERPRETATION: The reported data demonstrate that the use of the microRNA-based classifier in the real-world can reduce the rate of thyroid surgeries with robust performance and support clinical decision-making. FUNDING: The São Paulo Research-Foundation (FAPESP) and Onkos.

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OBJECTIVE: To compare the diagnosis of bacterial vaginosis (BV) by Nugent scoring criteria (Nugent-BV) and the diagnosis of BV and/or aerobic vaginitis (AV) using Donders criteria (Donders-BV/AV) for identifying Molecular-BV detected by bacterial 16s rRNA profiling. METHODS: We enrolled 512 women of reproductive age in Brazil with data available on Nugent and Donders microscopic analysis and 16S rRNA sequencing. We constructed receiver operating characteristic (ROC) curves of Nugent-BV and Donders-BV/AV and calculated their area under the curves (AUCs) and 95% confidence interval (CI) for matching Molecular-BV. RESULTS: A total of 155 (28.7%) participants were positive for Nugent-BV. Donders-BV and -AV were detected in 90 (17.6%) and 75 (14.6%) participants, respectively, while 28 (5.5%) had concurrent Donders-BV and -AV. Molecular-BV was identified in 139 (27.1%) participants. Analysis of ROC curves showed that diagnosis of Nugent-BV more accurately aligned with presence of Molecular-BV (AUC: 0.88, 95% CI: 0.84-0.91) when compared to Donders-AV/BV (AUC: 0.84; CI: 0.80-0.87) (P = 0.005). CONCLUSION: The use of Nugent-BV is more representative of Molecular-BV than Donders-AV/BV.

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The emergence of the COVID-19 pandemic prompted fast development of novel diagnostic methods of the etiologic virus SARS-CoV-2. Methods based on CRISPR-Cas systems have been particularly promising because they can achieve a similar sensitivity and specificity to the benchmark RT-qPCR, especially when coupled to an isothermal pre-amplification step. Furthermore, they have also solved inherent limitations of RT-qPCR that impede its decentralized use and deployment in the field, such as the need for expensive equipment, high cost per reaction, and delivery of results in hours, among others. In this review, we evaluate publicly available methods to detect SARS-CoV-2 that are based on CRISPR-Cas and isothermal amplification. We critically analyze the steps required to obtain a successful result from clinical samples and pinpoint key experimental conditions and parameters that could be optimized or modified to improve clinical and analytical outputs. The COVID outbreak has propelled intensive research in a short time, which is paving the way to develop effective and very promising CRISPR-Cas systems for the precise detection of SARS-CoV-2. This review could also serve as an introductory guide to new labs delving into this technology.

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The Curtobacterium genus is a member of the family Microbacteriaceae, and Curtobacterium species are recognized as plant pathogens. The aim of this study was to investigate a dubious result of species identification for an infection located on a catheter tip of a patient with Covid-19. A strain isolated from a catheter tip sample, identified by VITEK® 2 as Cronobacter spp., was submitted to polyphasic analysis: Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) using VITEK® MS, real-time polymerase chain reaction targeting dnaG gene, and 16S rRNA full gene Sanger sequencing analysis for confirmation. The strain presented negative result using qPCR and could not identified by MALDI-TOF MS. 16S rRNA full gene Sanger sequencing analysis identified the strain as Curtobacterium spp. The Gram-variable characteristic (Gram-negative instead of Gram-positive) of the isolated strain was the responsible for the misidentification by VITEK® 2 and VITEK® MS did not identify the strain. 16S rRNA full gene sequencing analysis identified the strain as Curtobacterium genus, but other complementary techniques are necessary to identify at species level.

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BACKGROUND: In the current COVID-19 pandemic, early and rapid diagnosis of potentially infected and contagious individuals enables containment of the disease through quarantine and contact tracing. The rapid global expansion of these diagnostic testing services raises questions concerning the current state of the art with regard to standardization of testing and quality assessment practices. The aim of this study was to provide a global overview of the test methods, laboratory procedures and quality assessment practices used for SARS-CoV-2 diagnostics. METHODS: The Molecular Diagnostics Committee of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC C-MD) initiated a survey among international laboratories performing molecular genetic detection of SARS-CoV-2. Questions on quality assurance, variant testing, sequencing and the transmission of findings were included in the survey. RESULTS: A total of 273 laboratories from 49 countries participated in the survey. The majority of the participating laboratories (92.2%) use reverse transcriptase polymerase chain reaction (RT-PCR). The majority of participating laboratories do not conduct testing to identify SARS CoV-2 variants. Participation in external quality assessment programs was reported by the majority of laboratories, however, 33.2% of the laboratories reported not participating in external quality assurance programmes. CONCLUSIONS: Based on the survey, molecular diagnostic methods for SARS-CoV-2 detection are clearly not standardized across different countries and laboratories. The survey found an array of responses in regard to sample preparation, collection, processing and reporting of results. This work suggests quality assurance is insufficiently performed by diagnostic laboratories conducting SARS-CoV-2 testing.

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OBJECTIVE: To determine the prevalence of Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) in the homeless population in Medellín, Colombia, using molecular diagnostic methods. It also intended to develop a demographic profile, exploring associated factors and the dynamics of the social and sexual interactions of this community. DESIGN: Cross-sectional study. SETTING: Two homeless care centres in Medellín, Colombia. PARTICIPANTS: Homeless individuals that assisted to the main homeless care centres of Medellín, Colombia from 2017 to 2019. PRIMARY AND SECONDARY OUTCOME MEASURES: The prevalence of CT and NG in this population using qPCR detection, factors associated with CT and NG infection, and the sociodemographic profile of the community. RESULTS: The prevalence of CT infection was 19.2%, while that of NG was 22.6%. Furthermore, being a female was significantly correlated to CT infection p<0.05 (adjusted OR, AOR 2.42, 95% CI 1.31 to 4.47). NG infection was significantly associated with factors such as: sexual intercourse while having a sexually transmitted infection p<0.05 (AOR 3.19, 95% CI 1.48 to 6.85), having more than 11 sexual partners in the last 6 months p=0.04 (AOR 2.91, 95% CI 1.04 to 8.09) and having daily intercourse p=0.05 (AOR 3.15, 95% CI 1.02 to 9.74). CONCLUSIONS: The prevalence of CT and NG was higher than that reported in the general population. Additionally, females had a higher percentage of infection compared with males.

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