Enhancing a SARS-CoV-2 nucleocapsid antigen test sensitivity with cost efficient strategy through a cotton intermembrane insertion.

Lateral flow antigen tests have been widely used in the Covid-19 pandemic, allowing faster diagnostic test results and preventing further viral spread through isolation of infected individuals. Accomplishment of this screening must be performed with tests that show satisfactory sensitivity in order to successfully detect the target protein and avoid false negatives. The aim of this study was to create a lateral flow test that could detect SARS-CoV-2 nucleocapsid protein in low concentrations that were comparable to the limits of detection claimed by existing tests from the market. To do so, several adjustments were necessary during research and development of the prototypes until they were consistent with these criteria. The proposed alternatives of increasing the test line antibody concentration and addition of an intermembrane between the conjugate pad and the nitrocellulose membrane were able to increase the sensitivity four-fold and generate a new rapid test prototype called "lateral flow intermembrane immunoassay test" (LFIIT). This prototype showed an adequate limit of detection (2.0 ng mL-1) while maintaining affordability and simplicity in manufacturing processes.

Development of an optimized colorimetric RT-LAMP for SARS-CoV-2 assay with enhanced procedure controls for remote diagnostics.

The coronavirus pandemic accentuated the need for molecular diagnostic tests. A technique highly used to this end is the Polymerase Chain Reaction (PCR)-a sensitive and specific technique commonly used as the gold standard for molecular diagnostics. However, it demands highly trained personnel and high-maintenance equipment and is relatively time-consuming. An alternative is the Loop-Mediated Isothermal Amplification (LAMP) technique, which doesn't need sample purification or expensive equipment, and is similar to PCR when compared in sensitivity and specificity. In this paper, we developed an optimized colorimetric Reverse Transcriptase Loop-Mediated Isothermal Amplification (RT-LAMP) Point-of-Care test using a portable device to diagnose COVID-19. Variables such as concentration of primers, magnesium sulfate, betaine, hydrochloride guanidine, Bst, and temperature of the reactions were tested. We also created a pipetting quality control system-using a combination of dyes-to avoid false negatives due to a lack of samples added to the reaction test tube. Mineral oil was incorporated in the composition of the RT-LAMP reactions to avoid evaporation when a heating lid isn't available. The final RT-LAMP test is tenfold more sensitive when compared to the WarmStart Colorimetric Master mix from New England Biolabs with a sensitivity of 5 copies per µL.

Hilab system, a new point-of-care hematology analyzer supported by the Internet of Things and Artificial Intelligence.

The complete blood count (CBC) is one of the most requested tests by physicians. CBC tests, most realized in conventional hematological analyzers, are restricted to centralized laboratories due to frequent maintenance, large devices, and expensive costs required. On the other hand, most handheld CBC devices commercially available show high prices and are not liable to calibration or control procedures, which results in poor quality compared to standard hematology instruments. The Hilab system is a small-handed hematological platform that uses microscopy and chromatography techniques for blood cells and hematimetric parameters analysis through artificial intelligence, machine learning, and deep learning techniques. For clinical evaluation of the handheld CBC device, 450 blood samples were analyzed. The samples encompassed normal (82%) and pathological conditions (18%), such as thalassemias (2.2%), anemias (6.6%), and infections (9.2%). For all analytes, accuracy, precision, method comparison, and flagging capabilities of the Hilab System, were compared with the Sysmex XE-2100 (Sysmex, Japan) results. The sample source (venous and capillary) influences were also evaluated. Pearson correlation, Student t test, bias, and the Bland-Altman plot of each blood count analyte were calculated and shown. The significance level was set at p ≤ 0.05. For clinical evaluation, Hilab System and the Sysmex XE-2100 showed a strong correlation (r ≥ 0.9) for most evaluated parameters. In the precision study, analytes showed CV inside the limits established according to European Federation of Clinical Chemistry and Laboratory Medicine guidelines. The flagging capabilities of the Hilab system, compared to the manual microscopy technique, presented high sensibility, specificity, and accuracy. Venous and capillary samples (p > 0.05) do not differ statistically. Considering the need for point-of-care CBCs, the study indicated that the Hilab system provides fast, accurate, low cost, and robust analysis for reliable clinical use.

The Experience of Implementing a National Antimicrobial Resistance Surveillance System in Brazil.

Antimicrobial resistance (AMR) is a major public health threat of global proportions, which has the potential to lead to approximately ten million deaths per year by 2050. Pressured by this wicked problem, in 2014, the World Health Organization launched a call for member states to share AMR data through the implementation of the Global Antimicrobial Resistance Surveillance System (GLASS), to appropriately scale and monitor the general situation world-widely. In 2017, Brazil joined GLASS and, in 2018, started its own national antimicrobial surveillance program (BR-GLASS) to understand the impact of resistance in the country. We compiled data obtained from the complete routine of three hospitals' microbiology labs during the year of 2018. This pilot data sums up to 200,874 antimicrobial susceptibility test results from 11,347 isolates. It represents 119 different microorganisms recovered from 44 distinct types of clinical samples. Specimens came from patients originating from 301 Brazilian cities, with 4,950 of these isolates from presumed Healthcare-Associated Infections (HAIs) and the other 6,397 community-acquired cases. The female population offered 58% of the collected samples, while the other 42% were of male origin. The urinary tract was the most common topography (6,372/11,347 isolates), followed by blood samples (2,072/11,347). Gram-negative predominated the bacterial isolates: Escherichia coli was the most prevalent in general, representing 4,030 isolates (89.0% of these from the urinary tract). Coagulase-negative Staphylococci were the most prevalent bacteria in blood samples. Besides these two species, the ESKAPE group have consolidated their prevalence. Regarding drug susceptibility results, 141,648 (70.5%) were susceptible, 9,950 (4.9%) intermediate, and 49,276 (24.5%) resistant. Acinetobacter baumannii was the most worrisome microorganism, with 65.3% of the overall antimicrobial susceptibility tests showing resistance, followed by ESBL-producing Klebsiella pneumoniae, with a global resistance rate of 59%. Although this is a pilot project (still limited to one state), this database shows the importance of a nation-wide surveillance program,[153mm][-12mm] Q14 especially considering it already had patients coming from 301 distinct counties and 18 different states. The BR-GLASS Program is an ongoing project that intends to encompass at least 95 hospitals distributed in all five geographical regions in Brazil within the next 5 years.